scholarly journals Isolation of Bacteroides fragilis Mutants with In Vivo Growth Defects by Using Tn4400′, a Modified Tn4400 Transposition System, and a New Screening Method

2000 ◽  
Vol 68 (1) ◽  
pp. 415-419 ◽  
Author(s):  
Yixin P. Tang ◽  
Michael H. Malamy
Keyword(s):  
Tissue Culture ◽  
Screening Method ◽  
Bacteroides Fragilis ◽  
Normal Growth ◽  
Growth Characteristics ◽  
Rich Medium ◽  
Growth Defects ◽  
Mutant Strains ◽  
In Vivo Growth

ABSTRACT A modified version of the Bacteroides fragilistransposon Tn4400, designated Tn4400′, enabling rapid isolation and analysis of B. fragilis mutants has been constructed. To identify potential virulence factors, Tn4400′-generated mutants were screened by a new method; this resulted in the isolation of 21 mutant strains with impaired growth characteristics on tissue culture monolayers but normal growth in rich medium anaerobically.

scholarly journals Bone marrow from children in relapse with pre-B acute lymphoblastic leukemia proliferates and disseminates rapidly in scid mice

Blood ◽  
1991 ◽  
Vol 78 (11) ◽  
pp. 2973-2981 ◽  
Author(s):  
S Kamel-Reid ◽  
M Letarte ◽  
M Doedens ◽  
A Greaves ◽  
B Murdoch ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
Poor Prognosis ◽  
Lymphoblastic Leukemia ◽  
Growth Characteristics ◽  
Scid Mice ◽  
Leukemic Cells ◽  
Murine Bone Marrow ◽  
In Vivo Growth

Bone marrow samples from patients with pre-B acute lymphoblastic leukemia (pre-B ALL), either at diagnosis or at relapse, were transplanted into scid mice to determine whether these freshly obtained leukemic cells could proliferate in vivo and whether there were any differences in their in vivo growth characteristics. Cells from three patients who relapsed within 13 months of diagnosis proliferated rapidly in the murine bone marrow, spleen, and thymus, invaded peripheral organs, and resulted in morbidity and mortality of the animals within 4 to 16 weeks. Cells from two patients who relapsed 3.5 years after diagnosis grew much slower than the early relapse samples, taking up to 30 weeks to infiltrate the bone marrow of recipient mice. In contrast, leukemic cells were absent or were detected at low numbers in scid mice transplanted with cells obtained at diagnosis from three patients who have not yet relapsed. These results show an increased ability of leukemic cells from patients with aggressive lymphoblastic leukemia of poor prognosis to proliferate in scid mice.

scholarly journals Multiple knockout mouse models reveal lincRNAs are required for life and brain development

eLife ◽  
2013 ◽  
Vol 2 ◽  
Author(s):  
Martin Sauvageau ◽  
Loyal A Goff ◽  
Simona Lodato ◽  
Boyan Bonev ◽  
Abigail F Groff ◽  
...  
Keyword(s):  
Incomplete Penetrance ◽  
Functional Roles ◽  
Genetic Ablation ◽  
Physiological Conditions ◽  
Growth Defects ◽  
Initial Characterization ◽  
Mutant Strains ◽  
Functional Relevance ◽  
Functional Investigation

Many studies are uncovering functional roles for long noncoding RNAs (lncRNAs), yet few have been tested for in vivo relevance through genetic ablation in animal models. To investigate the functional relevance of lncRNAs in various physiological conditions, we have developed a collection of 18 lncRNA knockout strains in which the locus is maintained transcriptionally active. Initial characterization revealed peri- and postnatal lethal phenotypes in three mutant strains (Fendrr, Peril, and Mdgt), the latter two exhibiting incomplete penetrance and growth defects in survivors. We also report growth defects for two additional mutant strains (linc–Brn1b and linc–Pint). Further analysis revealed defects in lung, gastrointestinal tract, and heart in Fendrr−/− neonates, whereas linc–Brn1b−/− mutants displayed distinct abnormalities in the generation of upper layer II–IV neurons in the neocortex. This study demonstrates that lncRNAs play critical roles in vivo and provides a framework and impetus for future larger-scale functional investigation into the roles of lncRNA molecules.

scholarly journals The Ribosome-Bound Chaperones RAC and Ssb1/2p Are Required for Accurate Translation in Saccharomyces cerevisiae

2004 ◽  
Vol 24 (20) ◽  
pp. 9186-9197 ◽  
Author(s):  
Magdalena Rakwalska ◽  
Sabine Rospert
Keyword(s):  
Translation Termination ◽  
Ribosomal Subunit ◽  
Reporter System ◽  
Translational Fidelity ◽  
Growth Defects ◽  
Mutant Strains ◽  
Translation Termination Factor ◽  
Combined Data

ABSTRACT The chaperone homologs RAC (ribosome-associated complex) and Ssb1/2p are anchored to ribosomes; Ssb1/2p directly interacts with nascent polypeptides. The absence of RAC or Ssb1/2p results in a similar set of phenotypes, including hypersensitivity against the aminoglycoside paromomycin, which binds to the small ribosomal subunit and compromises the fidelity of translation. In order to understand this phenomenon we measured the frequency of translation termination and misincorporation in vivo and in vitro with a novel reporter system. Translational fidelity was impaired in the absence of functional RAC or Ssb1/2p, and the effect was further enhanced by paromomycin. The mutant strains suffered primarily from a defect in translation termination, while misincorporation was compromised to a lesser extent. Consistently, a low level of soluble translation termination factor Sup35p enhanced growth defects in the mutant strains. Based on the combined data we conclude that RAC and Ssb1/2p are crucial in maintaining translational fidelity beyond their postulated role as chaperones for nascent polypeptides.

Capsulated cells of Aeromonas salmonicida grown in vitro have different functional properties than capsulated cells grown in vivo

1995 ◽  
Vol 41 (10) ◽  
pp. 941-945 ◽  
Author(s):  
Rafael A. Garduño ◽  
William W. Kay
Keyword(s):  
Functional Properties ◽  
Surface Antigens ◽  
Aeromonas Salmonicida ◽  
Serum Resistance ◽  
Rich Medium ◽  
Increased Sensitivity ◽  
In Vivo Growth ◽  
Capsular Material

When grown in vivo in the peritoneal cavity of rainbow trout, Aeromonas salmonicida produces a clearly defined capsule with virulence-related functions. Aeromonas salmonicida grown in vitro in a glucose-rich medium (GRM) has also been reported to produce capsular material. Because in vitro mimicry of in vivo induced traits is highly desirable in vaccine design, the extent to which growth in GRM mimicked in vivo growth was examined. Antibodies specific to in vivo grown cells partially labeled the surface of GRM-grown cells, as well as two distinct proteins (81 700 and 41 000 Mr) in immunoblots of mutants with S-layer or lipopolysaccharide defects. GRM-grown strains showed an increased sensitivity to trout serum in contradistinction to the complete serum resistance of in vivo grown cells; as well, GRM-grown cells were more adherent to trout macrophages. Thus in spite of possessing some surface antigens normally expressed in vivo, cells grown on solid GRM did not possess all functional properties of in vivo grown cells.Key words: Aeromonas salmonicida, bacterial capsules, mimicry of in vivo conditions, furunculosis vaccines.

In vivo growth characteristics of the Landschütz ascites tumour

1978 ◽  
Vol 147 (1) ◽  
pp. 134-139 ◽  
Author(s):  
John Brazil ◽  
Hugh McLaughlin
Keyword(s):  
Growth Characteristics ◽  
Ascites Tumour ◽  
In Vivo Growth

scholarly journals Human Immunodeficiency Virus Bearing a Disrupted Central DNA Flap Is Pathogenic In Vivo

2007 ◽  
Vol 81 (11) ◽  
pp. 6146-6150 ◽  
Author(s):  
Matthew D. Marsden ◽  
Jerome A. Zack
Keyword(s):  
Human Immunodeficiency Virus ◽  
Growth Characteristics ◽  
Deficient Mutant ◽  
Wild Type ◽  
Hiv Replication ◽  
Immunodeficiency Virus ◽  
In Vivo Growth ◽  
Kinetics Of

ABSTRACT The central DNA flap is an important component of lentiviral vectors, but its significance in the context of wild-type human immunodeficiency virus (HIV) is currently unclear. To address this issue, we have compared the in vitro infection kinetics of NL4-3 with those of a flap-deficient mutant and evaluated the in vivo growth characteristics of these viruses by using the SCID-hu mouse model of HIV infection. Flap-deficient virus was only modestly attenuated in vitro, as assessed by single-round and spreading infection assays, and exhibited levels of replication and pathogenesis close to those of the wild-type in vivo. Hence, an intact central flap is not essential for HIV replication.

scholarly journals Characterization of Bacteroides fragilis Hemolysins and Regulation and Synergistic Interactions of HlyA and HlyB

2006 ◽  
Vol 74 (4) ◽  
pp. 2304-2316 ◽  
Author(s):  
Kirstin P. Robertson ◽  
C. Jeffrey Smith ◽  
Andrea M. Gough ◽  
Edson R. Rocha
Keyword(s):  
Hemolytic Activity ◽  
Blot Hybridization ◽  
Bacteroides Fragilis ◽  
Blood Agar ◽  
Northern Blot Hybridization ◽  
Rich Medium ◽  
Independent Manner ◽  
Hemolytic Assay ◽  
Growth Deficiency

ABSTRACT This study describes the presence of 10 hemolysin orthologs in the genome of the opportunistic human anaerobic pathogen Bacteroides fragilis, which is currently classified as a nonhemolytic bacterium. The hemolysins were designated HlyA through HlyI plus HlyIII. All cloned hemolysin genes were able to confer hemolytic activity to a nonhemolytic Escherichia coli strain on blood agar plates. Interestingly, HlyH was found to be present in the genome of the B. fragilis NCTC9343 strain but absent in strains 638R, YCH46, and Bacteroides thetaiotaomicron VPI-5482. The hemolysins HlyA, HlyB, and HlyIII were selected for further characterization. HlyA, HlyB, and HlyIII were cytolytic to erythrocytes on liquid hemolytic assay. When hlyA and hlyB were expressed together in a nonhemolytic E. coli strain, the strain showed enhanced hemolytic activity on blood agar plates. Further analysis revealed that HlyA and HlyB have synergistic hemolytic activity as detected by the liquid hemolytic assay. In addition, the two-component hemolysins HlyA and HlyB form a protein-protein complex in vivo as determined by bacterial two-hybrid system assay. The hlyB and hlyA genes are organized in an operon that is coordinately regulated by iron and oxygen. Northern blot hybridization analysis revealed that hlyBA were expressed as a bicistronic mRNA induced approximately 2.5-fold under low-iron conditions and repressed in iron-rich medium. The normal iron-regulated expression of hlyBA mRNA was lost in the furA mutant strain. In contrast, the hlyA gene was also expressed as a single mRNA in iron-rich medium, but its expression was reduced approximately threefold under low-iron conditions in a Fur-independent manner. This suggests that hlyA alone is regulated by an unidentified iron-dependent regulator. Moreover, the expression levels of hlyBA and hlyA were reduced about threefold following oxygen exposure and treatment with hydrogen peroxide. Taken together, these results suggest that iron and oxidative stress have an effect on the control of hlyBA and hlyA transcriptional levels. A hlyBA mutant was constructed, and its hemolytic activity was greatly diminished compared to those of the hlyIII mutant and parent strains. In addition, the hlyBA mutant had a significant modification in colony morphology and growth deficiency compared to the parent strain. The implications of these findings for the pathophysiology of B. fragilis in extraintestinal infections and competition in ecological systems for this organism are discussed.

scholarly journals Bone marrow from children in relapse with pre-B acute lymphoblastic leukemia proliferates and disseminates rapidly in scid mice

Blood ◽  
1991 ◽  
Vol 78 (11) ◽  
pp. 2973-2981 ◽  
Author(s):  
S Kamel-Reid ◽  
M Letarte ◽  
M Doedens ◽  
A Greaves ◽  
B Murdoch ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
Poor Prognosis ◽  
Lymphoblastic Leukemia ◽  
Growth Characteristics ◽  
Scid Mice ◽  
Leukemic Cells ◽  
Murine Bone Marrow ◽  
In Vivo Growth

Abstract Bone marrow samples from patients with pre-B acute lymphoblastic leukemia (pre-B ALL), either at diagnosis or at relapse, were transplanted into scid mice to determine whether these freshly obtained leukemic cells could proliferate in vivo and whether there were any differences in their in vivo growth characteristics. Cells from three patients who relapsed within 13 months of diagnosis proliferated rapidly in the murine bone marrow, spleen, and thymus, invaded peripheral organs, and resulted in morbidity and mortality of the animals within 4 to 16 weeks. Cells from two patients who relapsed 3.5 years after diagnosis grew much slower than the early relapse samples, taking up to 30 weeks to infiltrate the bone marrow of recipient mice. In contrast, leukemic cells were absent or were detected at low numbers in scid mice transplanted with cells obtained at diagnosis from three patients who have not yet relapsed. These results show an increased ability of leukemic cells from patients with aggressive lymphoblastic leukemia of poor prognosis to proliferate in scid mice.

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