bluuebunny/crossref_metadata_2025_split huggingface dataset:
Functional Analysis of the
Lactococcus lactis galU
and
galE
Genes and Their Impact on Sugar Nucleotide and Exopolysaccharide Biosynthesis
Authors: Boels Ingeborg C. , Ramos Ana , Kleerebezem Michiel , de Vos Willem M. 
Published: July 2002

ABSTRACT

We studied the UDP-glucose pyrophosphorylase (
galU
) and UDP-galactose epimerase (
galE
) genes of
Lactococcus lactis
MG1363 to investigate their involvement in biosynthesis of UDP-glucose and UDP-galactose, which are precursors of glucose- and galactose-containing exopolysaccharides (EPS) in
L. lactis
. The lactococcal
galU
gene was identified by a PCR approach using degenerate primers and was found by Northern blot analysis to be transcribed in a monocistronic RNA. The
L. lactis galU
gene could complement an
Escherichia coli galU
mutant, and overexpression of this gene in
L. lactis
under control of the inducible
nisA
promoter resulted in a 20-fold increase in GalU activity. Remarkably, this resulted in approximately eightfold increases in the levels of both UDP-glucose and UDP-galactose. This indicated that the endogenous GalE activity is not limiting and that the GalU activity level in wild-type cells controls the biosynthesis of intracellular UDP-glucose and UDP-galactose. The increased GalU activity did not significantly increase NIZO B40 EPS production. Disruption of the
galE
gene resulted in poor growth, undetectable intracellular levels of UDP-galactose, and elimination of EPS production in strain NIZO B40 when cells were grown in media with glucose as the sole carbon source. Addition of galactose restored wild-type growth in the
galE
disruption mutant, while the level of EPS production was approximately one-half the wild-type level.
Spatial and Resource Factors Influencing High Microbial Diversity in Soil
Authors: Zhou Jizhong , Xia Beicheng , Treves David S. , Wu L.-Y. , Marsh Terry L. , O’Neill Robert V. , Palumbo Anthony V. , Tiedje James M. 
Published: July 2002

ABSTRACT

To begin defining the key determinants that drive microbial community structure in soil, we examined 29 soil samples from four geographically distinct locations taken from the surface, vadose zone, and saturated subsurface using a small-subunit rRNA-based cloning approach. While microbial communities in low-carbon, saturated, subsurface soils showed dominance, microbial communities in low-carbon surface soils showed remarkably uniform distributions, and all species were equally abundant. Two diversity indices, the reciprocal of Simpson’s index (1/
D
) and the log series index, effectively distinguished between the dominant and uniform diversity patterns. For example, the uniform profiles characteristic of the surface communities had diversity index values that were 2 to 3 orders of magnitude greater than those for the high-dominance, saturated, subsurface communities. In a site richer in organic carbon, microbial communities consistently exhibited the uniform distribution pattern regardless of soil water content and depth. The uniform distribution implies that competition does not shape the structure of these microbial communities. Theoretical studies based on mathematical modeling suggested that spatial isolation could limit competition in surface soils, thereby supporting the high diversity and a uniform community structure. Carbon resource heterogeneity may explain the uniform diversity patterns observed in the high-carbon samples even in the saturated zone. Very high levels of chromium contamination (e.g., >20%) in the high-organic-matter soils did not greatly reduce the diversity. Understanding mechanisms that may control community structure, such as spatial isolation, has important implications for preservation of biodiversity, management of microbial communities for bioremediation, biocontrol of root diseases, and improved soil fertility.
Resolution of
Prochlorococcus
and
Synechococcus
Ecotypes by Using 16S-23S Ribosomal DNA Internal Transcribed Spacer Sequences
Authors: Rocap Gabrielle , Distel Daniel L. , Waterbury John B. , Chisholm Sallie W. 
Published: July 2002

ABSTRACT

Cultured isolates of the marine cyanobacteria
Prochlorococcus
and
Synechococcus
vary widely in their pigment compositions and growth responses to light and nutrients, yet show greater than 96% identity in their 16S ribosomal DNA (rDNA) sequences. In order to better define the genetic variation that accompanies their physiological diversity, sequences for the 16S-23S rDNA internal transcribed spacer (ITS) region were determined in 32
Prochlorococcus
isolates and 25
Synechococcus
isolates from around the globe. Each strain examined yielded one ITS sequence that contained two tRNA genes. Dramatic variations in the length and G+C content of the spacer were observed among the strains, particularly among
Prochlorococcus
strains. Secondary-structure models of the ITS were predicted in order to facilitate alignment of the sequences for phylogenetic analyses. The previously observed division of
Prochlorococcus
into two ecotypes (called high and low-B/A after their differences in chlorophyll content) were supported, as was the subdivision of the high-B/A ecotype into four genetically distinct clades. ITS-based phylogenies partitioned marine cluster A
Synechococcus
into six clades, three of which can be associated with a particular phenotype (motility, chromatic adaptation, and lack of phycourobilin). The pattern of sequence divergence within and between clades is suggestive of a mode of evolution driven by adaptive sweeps and implies that each clade represents an ecologically distinct population. Furthermore, many of the clades consist of strains isolated from disparate regions of the world's oceans, implying that they are geographically widely distributed. These results provide further evidence that natural populations of
Prochlorococcus
and
Synechococcus
consist of multiple coexisting ecotypes, genetically closely related but physiologically distinct, which may vary in relative abundance with changing environmental conditions.
Role of
Bacillus thuringiensis
Cry1 δ Endotoxin Binding in Determining Potency during Lepidopteran Larval Development
Authors: Gilliland Androulla , Chambers Catherine E. , Bone Eileen J. , Ellar David J. 
Published: July 2002

ABSTRACT

Five economically important crop pests,
Manduca sexta
,
Pieris brassicae
,
Mamestra brassicae
,
Spodoptera exigua
, and
Agrotis ipsilon
, were tested at two stages of larval development for susceptibility to
Bacillus thuringiensis
toxins Cry1Ac, Cry1Ca, Cry1J, and Cry1Ba. Bioassay results for
M. sexta
showed that resistance to all four Cry toxins increased from the neonate stage to the third-instar stage; the increase in resistance was most dramatic for Cry1Ac, the potency of which decreased 37-fold. More subtle increases in resistance during larval development were seen in
M. brassicae
for Cry1Ca and in
P. brassicae
for Cry1Ac and Cry1J. By contrast, the sensitivity of
S. exigua
did not change during development. At both larval stages,
A. ipsilon
was resistant to all four toxins. Because aminopeptidase N (APN) is a putative Cry1 toxin binding protein, APN activity was measured in neonate and third-instar brush border membrane vesicles (BBMV). With the exception of
S. exigua
, APN activity was found to be significantly lower in neonates than in third-instar larvae and thus inversely correlated with increased resistance during larval development. The binding characteristics of iodinated Cry1 toxins were determined for neonate and third-instar BBMV. In
M. sexta
, the increased resistance to Cry1Ac and Cry1Ba during larval development was positively correlated with fewer binding sites in third-instar BBMV than in neonate BBMV. The other species-instar-toxin combinations did not reveal positive correlations between potency and binding characteristics. The correlation between binding and potency was inconsistent for the species-instar-toxin combinations used in this study, reaffirming the complex mode of action of Cry1 toxins.
Improved Culturability of Soil Bacteria and Isolation in Pure Culture of Novel Members of the Divisions
Acidobacteria
,
Actinobacteria
,
Proteobacteria
, and
Verrucomicrobia
Authors: Janssen Peter H. , Yates Penelope S. , Grinton Bronwyn E. , Taylor Paul M. , Sait Michelle 
Published: July 2002

ABSTRACT

The culturability of bacteria in the bulk soil of an Australian pasture was investigated by using nutrient broth at 1/100 of its normal concentration (dilute nutrient broth [DNB]) as the growth medium. Three-tube most-probable-number serial dilution culture resulted in a mean viable count that was only 1.4% of the mean microscopically determined total cell count. Plate counts with DNB solidified with agar and with gellan gum resulted in viable counts that were 5.2 and 7.5% of the mean microscopically determined total cell count, respectively. Prior homogenization of the soil sample with an ultrasonic probe increased the viable count obtained by using DNB solidified with gellan gum to 14.1% of the mean microscopically determined cell count. A microscopic examination of the cell aggregates that remained after sonication revealed that the potential CFU count was only 70.4% of the total cell count, due to cells occurring as pairs or in clumps of three or more cells. Staining with SYTO 9 plus propidium iodide indicated that 91.3% of the cells in sonicated soil samples were potentially viable. Together, these findings suggest that the maximum achievable CFU count may be as low as 64.3% of the total cell count. Thirty isolates obtained from plate counting experiments performed with DNB as the growth medium were identified by comparative analysis of partial 16S rRNA gene sequences. A large proportion of these isolates represent the first known isolates of globally distributed groups of soil bacteria belonging to novel lineages within the divisions
Actinobacteria
,
Acidobacteria
,
Proteobacteria
, and
Verrucomicrobia
.
Prevalence of
Enterocytozoon bieneusi
in Swine: an 18-Month Survey at a Slaughterhouse in Massachusetts
Authors: Buckholt Michael A. , Lee John H. , Tzipori Saul 
Published: July 2002

ABSTRACT

Slaughterhouse pig samples were analyzed by PCR for
Enterocytozoon bieneusi
infection. Thirty-two percent were found to be positive, with rates being higher over the summer months. Three isolates from pigs were identical in their ribosomal internal transcribed spacer sequence to human
E. bieneusi
type D, two were identical to type F (from a pig), and nine were previously unreported. The viability of these spores was demonstrated by their ability to infect gnotobiotic piglets. The presence of the infection in liver was shown by in situ hybridization.
Novel Approach for Specific Detection of Herpes Simplex Virus Type 1 and 2 Antibodies and Immunoglobulin G and M Antibodies
Authors: Ohana Bella , Lipson Mark , Vered Noga , Srugo Isaac , Ahdut Moshe , Morag Abraham 
Published: July 2002

ABSTRACT
Recently a few new herpes simplex virus (HSV) type-specific serological diagnostic tests have been introduced to the commercial market, but these tests have some limitations. Moreover, it is not yet clear which commercial test can be regarded as a “gold standard” for the serodiagnosis of HSV infections. In order to improve the clinical diagnostic value of serological tests for the detection of HSV infections, we developed novel, competition-based enzyme-linked immunosorbent assays for the specific determination of HSV type 2 antibodies (SeroHSV2) and HSV type 1 antibodies (SeroHSV1) and two complementary tests for the detection of HSV immunoglobulin M (IgM) and IgG antibodies (SeroHSV IgM and SeroHSV IgG). These four new kits were evaluated in comparison with some commercial kits for the detection of HSV antibodies that are commonly used at present in Israeli clinical laboratories. The results indicate that SeroHSV2 is highly sensitive (>92%) and highly specific (>94%). SeroHSV2 does not cross-react with other alphaherpesvirus antibodies. SeroHSV1 is highly sensitive (>94%) and specific (>91%) compared to four commercial available kits. SeroHSV IgM is highly specific (>92%) in comparison with other commercial HSV IgM tests. The sensitivity of SeroHSV IgM ranges between 50 and 70% compared to these tests. Further investigation of the discrepant results obtained by using in-house competition tests indicated that SeroHSV IgM is more sensitive. SeroHSV IgG was also found to be highly sensitive (>94%) and highly specific (>92%) compared to the other commercial HSV IgG tests.
Antibody Responses to MAP 1B and Other
Cowdria ruminantium
Antigens Are Down Regulated in Cattle Challenged with Tick-Transmitted Heartwater
Authors: Semu S. M. , Peter T. F. , Mukwedeya D. , Barbet A. F. , Jongejan F. , Mahan S. M. 
Published: July 2002

ABSTRACT

Serological diagnosis of heartwater or
Cowdria ruminantium
infection has been hampered by severe cross-reactions with antibody responses to related ehrlichial agents. A MAP 1B indirect enzyme-linked immunosorbent assay that has an improved specificity and sensitivity for detection of immunoglobulin G (IgG) antibodies has been developed to overcome this constraint (A. H. M. van Vliet, B. A. M. Van der Zeijst, E. Camus, S. M. Mahan, D. Martinez, and F. Jongejan, J. Clin. Microbiol. 33:2405–2410, 1995). When sera were tested from cattle in areas of endemic heartwater infection in Zimbabwe, only 33% of the samples tested positive in this assay despite a high infection pressure (S. M. Mahan, S. M. Samu, T. F. Peter, and F. Jongejan, Ann. N.Y. Acad. Sci 849:85–87, 1998). To determine underlying causes for this observation, the kinetics of MAP 1B-specific IgG antibodies in cattle after tick-transmitted
C. ruminantium
infection and following recovery were investigated. Sera collected weekly over a period of 52 weeks from 37 cattle, which were naturally or experimentally infected with
C. ruminantium
via
Amblyomma hebraeum
ticks, were analyzed. MAP 1B-specific IgG antibody responses developed with similar kinetics in both field- and laboratory-infected cattle. IgG levels peaked at 4 to 9 weeks after tick infestation and declined to baseline levels between 14 and 33 weeks, despite repeated exposure to infected ticks and the establishment of a carrier state as demonstrated by PCR and xenodiagnosis. Some of the serum samples from laboratory, and field-infected cattle were also analyzed by immunoblotting and an indirect fluorescent-antibody test (IFAT) to determine whether this observed seroreversion was specific to the MAP 1B antigen. Reciprocal IFAT and immunoblot MAP 1-specific antibody titres peaked at 5 to 9 weeks after tick infestation but also declined between 30 and 45 weeks. This suggests that MAP 1B-specific IgG antibody responses and antibody responses to other
C. ruminantium
antigens are down regulated in cattle despite repeated exposure to
C. ruminantium
via ticks. Significantly, serological responses to the MAP 1B antigen may not be a reliable indicator of
C. ruminantium
exposure in cattle in areas of endemic heartwater infection.
Vγ9Vδ2 T Cells in Human Legionellosis
Authors: Kroca Michal , Johansson Anders , Sjöstedt Anders , Tärnvik Arne 
Published: July 2002

ABSTRACTIn humans, expansion of circulating Vγ9Vδ2 T cells seems to be a pathophysiological denominator shared by protozoan and intracellular bacterial diseases. The assumption was tested here on legionellosis, a condition conforming to the category but not yet described with respect to γδ T cells. Levels of Vγ9Vδ2 T cells in peripheral blood were measured at various intervals in 14 subjects undergoing a Pontiac fever-like disease, shown by serological investigation to be caused byLegionella micdadei. In samples obtained 4 to 6 days after the onset of the disease, the mean percentage (± the standard deviation) of Vγ9Vδ2+T cells among CD3+cells was 1.0% ± 0.5%, compared to 5.0% ± 3.9% in healthy control subjects (P< 0.001). Thereafter, a pronounced increase occurred and at 2 to 7 weeks after onset, mean peak levels were as high as ≈15%. During the next 6 months, values slowly declined, although without reaching the normal range. Percentages of γδ+T cells expressing tumor necrosis factor alpha or gamma interferon in response to phorbol myristate acetate were assayed in vitro. At 14 to 16 days after the onset of disease, the expression of both cytokines was increased (P< 0.01), whereas at 5 to 7 weeks, the expression of tumor necrosis factor alpha was decreased (P< 0.05), possibly reflecting modulation of an inflammatory response. In conclusion, Pontiac fever was found to be associated with a pronounced and long-lasting expansion of Vγ9Vδ2 T cells, implying that the subset may also be pathophysiologically important in a mild and transient form of intracellular bacterial diseases. Surprisingly, the expansion was preceded by a depletion of circulatory Vγ9Vδ2 T cells. Possibly, Vγ9Vδ2 T cells are initially recruited to a site of infection before they expand in response to antigen and occur in high numbers in blood.
Leptospirosis
Authors: Levett Paul N. 
Published: July 2002

SUMMARYLeptospirosis is a worldwide zoonotic infection with a much greater incidence in tropical regions and has now been identified as one of the emerging infectious diseases. The epidemiology of leptospirosis has been modified by changes in animal husbandry, climate, and human behavior. Resurgent interest in leptospirosis has resulted from large outbreaks that have received significant publicity. The development of simpler, rapid assays for diagnosis has been based largely on the recognition that early initiation of antibiotic therapy is important in acute disease but also on the need for assays which can be used more widely. In this review, the complex taxonomy of leptospires, previously based on serology and recently modified by a genotypic classification, is discussed, and the clinical and epidemiological value of molecular diagnosis and typing is also evaluated.
Heterologously Expressed
Staphylococcus aureus
Fibronectin-Binding Proteins Are Sufficient for Invasion of Host Cells
Authors: Sinha Bhanu , Francois Patrice , Que Yok-Ai , Hussain Muzaffar , Heilmann Christine , Moreillon Philippe , Lew Daniel , Krause Karl-Heinz , Peters Georg , Herrmann Mathias 
Published: July 2002

ABSTRACT

Staphylococcus aureus
invasion of mammalian cells, including epithelial, endothelial, and fibroblastic cells, critically depends on fibronectin bridging between
S. aureus
fibronectin-binding proteins (FnBPs) and the host fibronectin receptor integrin α
5
β
1
(B. Sinha et al., Cell. Microbiol. 1:101–117, 1999). However, it is unknown whether this mechanism is sufficient for
S. aureus
invasion. To address this question, various
S. aureus
adhesins (FnBPA, FnBPB, and clumping factor [ClfA]) were expressed in
Staphylococcus carnosus
and
Lactococcus lactis
subsp.
cremoris
. Both noninvasive gram-positive microorganisms are genetically distinct from
S. aureus
, lack any known
S. aureus
surface protein, and do not bind fibronectin. Transformants of
S. carnosus
and
L. lactis
harboring plasmids coding for various
S. aureus
surface proteins (FnBPA, FnBPB, and ClfA) functionally expressed adhesins (as determined by bacterial clumping in plasma, specific latex agglutination, Western ligand blotting, and binding to immobilized and soluble fibronectin). FnBPA or FnBPB but not of ClfA conferred invasiveness to
S. carnosus
and
L. lactis
. Invasion of 293 cells by transformants was comparable to that of strongly invasive
S. aureus
strain Cowan 1. Binding of soluble and immobilized fibronectin paralleled invasiveness, demonstrating that the amount of accessible surface FnBPs is rate limiting. Thus,
S. aureus
FnBPs confer invasiveness to noninvasive, apathogenic gram-positive cocci. Furthermore, FnBP-coated polystyrene beads were internalized by 293 cells, demonstrating that FnBPs are sufficient for invasion of host cells without the need for (
S. aureus
-specific) coreceptors.
Expression of the Nitric Oxide Synthase 2 Gene Is Not Essential for Early Control ofMycobacterium tuberculosisin the Murine Lung
Authors: Cooper Andrea M. , Pearl John E. , Brooks Jason V. , Ehlers Stefan , Orme Ian M. 
Published: July 2002

ABSTRACTThe interleukin-12 and gamma interferon (IFN-γ) pathway of macrophage activation plays a pivotal role in controlling tuberculosis. In the murine model, the generation of supplementary nitric oxide by the induction of the nitric oxide synthase 2 (NOS2) gene product is considered the principal antimicrobial mechanism of IFN-γ-activated macrophages. Using a low-dose aerosol-mediated infection model in the mouse, we have investigated the role of nitric oxide in controllingMycobacterium tuberculosisin the lung. In contrast to the consequences of a systemic infection, a low dose of bacteria introduced directly into the lungs of mice lacking the NOS2 gene is controlled almost as well as in intact animals. This is in contrast to the rapid progression of disease in mice lacking IFN-γ or a key member of the IFN signaling pathway, interferon regulatory factor 1. Thus while IFN-γ is pivotal in early control of bacterial growth in the lung, this control does not completely depend upon the expression of the NOS2 gene. The absence of inducible nitric oxide in the lung does, however, result in increased polymorphonuclear cell involvement and eventual necrosis in the pulmonary granulomas of the infected mice lacking the NOS2 gene.
Optimizing the Germfree Mouse Model for In Vivo Evaluation of Oral
Vibrio cholerae
Vaccine and Vector Strains
Authors: Crean Thomas I. , John Manohar , Calderwood Stephen B. , Ryan Edward T. 
Published: July 2002

ABSTRACT

The germfree mouse model of
Vibrio cholerae
infection can be used to judge immune responses to
V. cholerae
vaccine and vector strains. In the original model, a single oral inoculation was administered on day 0, a booster oral inoculation was administered on day 14, and immune responses were analyzed with samples collected on day 28. Unfortunately, immune responses in this model frequently were low level, and interanimal variability occurred. In order to improve this model, we evaluated various primary and booster
V. cholerae
inoculation schedules. The most prominent systemic and mucosal antibody responses were measured in mice that received a multiple primary inoculation series on days 0, 2, 4, and 6 and booster inoculations on days 28 and 42. These modifications result in improved preliminary evaluation of
V. cholerae
vaccine and vector strains in mice.
Staphylococcal Exfoliative Toxins Cleave α- and β-Melanocyte-Stimulating Hormones
Authors: Rago James V. , Vath Gregory M. , Tripp Timothy J. , Bohach Gregory A. , Ohlendorf Douglas H. , Schlievert Patrick M. 
Published: July 2002

ABSTRACTThe staphylococcal exfoliative toxins (ETs) A and B (ETA and ETB) are 27-kDa exotoxins produced by certain strains ofStaphylococcus aureusand are the causative agents of staphylococcal scalded-skin syndrome. The crystal structures of the ETs strongly indicate that the proteins are members of the serine protease family of enzymes, although protease activity until now has not yet been conclusively demonstrated. Here, we show that the peptide β-melanocyte-stimulating hormone (β-MSH) is cleaved by ETA and that both ETA and ETB are capable of cleaving α-MSH. Both toxins exhibit cleavage at specific glutamic acid residues in MSH peptides. Moreover, biologically inactive mutants of ETA were incapable of cleaving β-MSH.