In vivo and in vitro phenotypic differences between Great Lakes VHSV genotype IVb isolates with sequence types vcG001 and vcG002

The highly similar cytoplasmic β- and γ-actins differ by only four functionally similar amino acids, yet previous in vitro and in vivo data suggest that they support unique functions due to striking phenotypic differences between Actb and Actg1 null mouse and cell models. To determine whether the four amino acid variances were responsible for the functional differences between cytoplasmic actins, we gene edited the endogenous mouse Actb locus to translate γ-actin protein. The resulting mice and primary embryonic fibroblasts completely lacked β-actin protein, but were viable and did not present with the most overt and severe cell and organismal phenotypes observed with gene knockout. Nonetheless, the edited mice exhibited progressive high-frequency hearing loss and degeneration of actin-based stereocilia as previously reported for hair cell-specific Actb knockout mice. Thus, β-actin protein is not required for general cellular functions, but is necessary to maintain auditory stereocilia.

Download Full-text

Cryptococcus neoformans Dual GDP-Mannose Transporters and Their Role in Biology and Virulence

Eukaryotic Cell ◽

10.1128/ec.00054-14 ◽

2014 ◽

Vol 13 (6) ◽

pp. 832-842 ◽

Cited By ~ 13

Author(s):

Zhuo A. Wang ◽

Cara L. Griffith ◽

Michael L. Skowyra ◽

Nichole Salinas ◽

Matthew Williams ◽

...

Keyword(s):

Cryptococcus Neoformans ◽

Protein Glycosylation ◽

Content Type ◽

Phenotypic Differences ◽

Major Virulence Factor ◽

Transmembrane Transporters ◽

Mutant Cells ◽

Made In

ABSTRACTCryptococcus neoformansis an opportunistic yeast responsible for lethal meningoencephalitis in humans. This pathogen elaborates a polysaccharide capsule, which is its major virulence factor. Mannose constitutes over one-half of the capsule mass and is also extensively utilized in cell wall synthesis and in glycosylation of proteins and lipids. The activated mannose donor for most biosynthetic reactions, GDP-mannose, is made in the cytosol, although it is primarily consumed in secretory organelles. This compartmentalization necessitates specific transmembrane transporters to make the donor available for glycan synthesis. We previously identified two cryptococcal GDP-mannose transporters, Gmt1 and Gmt2. Biochemical studies of each protein expressed inSaccharomyces cerevisiaeshowed that both are functional, with similar kinetics and substrate specificitiesin vitro. We have now examined these proteinsin vivoand demonstrate that cells lacking Gmt1 show significant phenotypic differences from those lacking Gmt2 in terms of growth, colony morphology, protein glycosylation, and capsule phenotypes. Some of these observations may be explained by differential expression of the two genes, but others suggest that the two proteins play overlapping but nonidentical roles in cryptococcal biology. Furthermore,gmt1 gmt2double mutant cells, which are unexpectedly viable, exhibit severe defects in capsule synthesis and protein glycosylation and are avirulent in mouse models of cryptococcosis.

Download Full-text

Divergent replication kinetics of two phenotypically different parvoviruses of rats

Journal of General Virology ◽

10.1099/0022-1317-82-3-537 ◽

2001 ◽

Vol 82 (3) ◽

pp. 537-546 ◽

Cited By ~ 3

Author(s):

Lisa J. Ball-Goodrich ◽

Elizabeth Johnson ◽

Robert Jacoby

Keyword(s):

Infectious Agent ◽

Protein Accumulation ◽

Sequence Comparisons ◽

Minute Virus Of Mice ◽

Phenotypic Differences ◽

Host Interactions ◽

Laboratory Rats ◽

Kinetics Of

Rat virus (RV) is an important infectious agent of laboratory rats because of its high prevalence and capacity to disrupt research. Additionally, RV infection serves as a model for characterizing virus–host interactions during acute, persistent and prenatal infection. Our research has examined the pathogenesis of two RV strains, RV-UMass and RV-Y. RV-UMass is more pathogenic, causes a higher level of persistent infection and transmits to the foetus after oronasal inoculation of the pregnant dam. To determine in vitro distinctions between the strains that may account for these differences and to provide a benchmark for characterizing virus replication in vivo, synchronized in vitro replication of both RV strains was defined and compared. The results demonstrated that RV replication has replicative intermediates, virus transcripts and proteins similar to those reported for the prototype parvovirus, minute virus of mice. However, the replicative cycle of RV-UMass was 12 h compared with 24 h for RV-Y, and RV-UMass and RV-Y differed in kinetics of virus DNA replication, transcription and protein accumulation. Additionally, in situ analysis correlated well with kinetics data as determined by Southern and Northern blot analysis. Sequence comparisons between the strains also determined coding differences that may contribute to phenotypic differences.

Download Full-text

Concentration-Dependent Selection of Small Phenotypic Differences in TEM β-Lactamase-Mediated Antibiotic Resistance

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.44.9.2485-2491.2000 ◽

2000 ◽

Vol 44 (9) ◽

pp. 2485-2491 ◽

Cited By ~ 89

Author(s):

Maria-Cristina Negri ◽

Marc Lipsitch ◽

Jesús Blázquez ◽

Bruce R. Levin ◽

Fernando Baquero

Keyword(s):

Amino Acid Replacement ◽

Single Amino Acid ◽

Chromosomal Marker ◽

Beta Lactamase ◽

Antibiotic Resistant ◽

Phenotypic Differences ◽

The Individual ◽

Selection Of

ABSTRACT In this paper, the first robust experimental evidence of in vitro and in vivo concentration-dependent selection of low-level antibiotic-resistant genetic variants is described. The work is based on the study of an asymmetric competition assay with pairs of isogenicEscherichia coli strains, differing only (apart from a neutral chromosomal marker) in a single amino acid replacement in a plasmid-mediated TEM-1 beta-lactamase enzyme, which results in the new TEM-12 beta-lactamase. The mixture was challenged by different antibiotic concentrations, both in vitro and in the animal model, and the selective process of the variant population was carefully monitored. A mathematical model was constructed to test the hypothesis that measured growth and killing rates of the individual TEM variants at different antibiotic concentrations could be used to predict quantitatively the strength of selection for TEM-12 observed in competition experiments at these different concentrations.

Download Full-text

Evaluation of an in vitro and in vivo model for experimental infection with Leishmania (Viannia) braziliensis and L. (V.) peruviana

Parasitology ◽

10.1017/s0031182007003848 ◽

2007 ◽

Vol 135 (3) ◽

pp. 319-326 ◽

Cited By ~ 16

Author(s):

D. GAMBOA ◽

K. TORRES ◽

S. DE DONCKER ◽

M. ZIMIC ◽

J. AREVALO ◽

...

Keyword(s):

Experimental Infection ◽

Parasite Species ◽

In Vivo Model ◽

Peruvian Andes ◽

Transmission Cycle ◽

Phenotypic Differences ◽

Enrich Culture ◽

Parasite Populations

SUMMARYLeishmania (Viannia) braziliensis and L. (V.) peruviana are two parasite species characterized by a very different pathogenicity in humans despite a high genetic similarity. We hypothesized previously that L. (V.) peruviana would descend from L. (V.) braziliensis and would have acquired its ‘peruviana’ character during the southward colonization and adaptation of the transmission cycle in the Peruvian Andes. In order to have a first appreciation of the differences in virulence between both species, we evaluated an in vitro and in vivo model for experimental infection. A procedure was adapted to enrich culture forms in infective stages and the purified metacyclics were used to infect macrophage cell lines and golden hamsters. The models were tested with 2 representative strains of L. (V.) braziliensis from cutaneous and mucosal origin respectively and 2 representative strains of L. (V.) peruviana from Northern and Southern Peru respectively. Our models were reproducible and sensitive enough to detect phenotypic differences among strains. We showed in vitro as well as in vivo that the L. (V.) braziliensis was more infective than L. (V.) peruviana. Furthermore, we found that in vitro infectivity patterns of the 4 strains analysed, were in agreement with the geographical structuring of parasite populations demonstrated in our previous studies. Further work is needed to confirm our results with more strains of different geographical origin and their specific clinical outcome. However, our data open new perspectives for understanding the process of speciation in Leishmania and its implications in terms of pathogenicity.

Download Full-text

Hammondia hammondi Harbors Functional Orthologs of the Host-Modulating Effectors GRA15 and ROP16 but Is Distinguished from Toxoplasma gondii by a Unique Transcriptional Profile

Eukaryotic Cell ◽

10.1128/ec.00215-14 ◽

2014 ◽

Vol 13 (12) ◽

pp. 1507-1518 ◽

Cited By ~ 10

Author(s):

Katelyn A. Walzer ◽

Gregory M. Wier ◽

Rachel A. Dam ◽

Ananth R. Srinivasan ◽

Adair L. Borges ◽

...

Keyword(s):

Toxoplasma Gondii ◽

Transcript Abundance ◽

Dense Granule ◽

Transcriptional Analysis ◽

Content Type ◽

Phenotypic Differences ◽

Functional Conservation ◽

High Transcript Abundance

ABSTRACTToxoplasma gondiiand its nearest extant relative,Hammondia hammondi, are phenotypically distinct despite their remarkable similarity in gene content, synteny, and functionality. To begin to identify genetic differences that might drive distinct infection phenotypes ofT. gondiiandH. hammondi, in the present study we (i) determined whether two known host-interacting proteins, dense granule protein 15 (GRA15) and rhoptry protein 16 (ROP16), were functionally conserved inH. hammondiand (ii) performed the first comparative transcriptional analysis ofH. hammondiandT. gondiisporulated oocysts. We found that GRA15 and ROP16 fromH. hammondi(HhGRA15 and HhROP16) modulate the host NF-κB and STAT6 pathways, respectively, when expressed heterologously inT. gondii. We also found the transcriptomes ofH. hammondiandT. gondiito be highly distinct. Consistent with the spontaneous conversion ofH. hammonditachyzoites into bradyzoites bothin vitroandin vivo,H. hammondihigh-abundance transcripts are enriched for genes that are of greater abundance inT. gondiibradyzoites. We also identified genes that are of high transcript abundance inH. hammondibut are poorly expressed in multipleT. gondiilife stages, suggesting that these genes are uniquely expressed inH. hammondi. Taken together, these data confirm the functional conservation of knownT. gondiivirulence effectors inH. hammondiand point to transcriptional differences as a potential source of the phenotypic differences between these species.

Download Full-text

Associations between Cryptococcus Genotypes, Phenotypes, and Clinical Parameters of Human Disease: A Review

Journal of Fungi ◽

10.3390/jof7040260 ◽

2021 ◽

Vol 7 (4) ◽

pp. 260

Author(s):

Marhiah C. Montoya ◽

Paul M. Magwene ◽

John R. Perfect

Keyword(s):

Fungal Pathogens ◽

Central Nervous System Disease ◽

Human Infection ◽

Phenotypic Differences ◽

System Disease ◽

Species Complexes ◽

Cryptococcal Disease ◽

In Vivo Animal Models

The genus Cryptococcus contains two primary species complexes that are significant opportunistic human fungal pathogens: C. neoformans and C. gattii. In humans, cryptococcosis can manifest in many ways, but most often results in either pulmonary or central nervous system disease. Patients with cryptococcosis can display a variety of symptoms on a spectrum of severity because of the interaction between yeast and host. The bulk of our knowledge regarding Cryptococcus and the mechanisms of disease stem from in vitro experiments and in vivo animal models that make a fair attempt, but do not recapitulate the conditions inside the human host. To better understand the dynamics of initiation and progression in cryptococcal disease, it is important to study the genetic and phenotypic differences in the context of human infection to identify the human and fungal risk factors that contribute to pathogenesis and poor clinical outcomes. In this review, we summarize the current understanding of the different clinical presentations and health outcomes that are associated with pathogenicity and virulence of cryptococcal strains with respect to specific genotypes and phenotypes.

Download Full-text

CD11cloB220+ interferon-producing killer dendritic cells are activated natural killer cells

Journal of Experimental Medicine ◽

10.1084/jem.20071451 ◽

2007 ◽

Vol 204 (11) ◽

pp. 2569-2578 ◽

Cited By ~ 112

Author(s):

Christian A.J. Vosshenrich ◽

Sarah Lesjean-Pottier ◽

Milena Hasan ◽

Odile Richard-Le Goff ◽

Erwan Corcuff ◽

...

Keyword(s):

Dendritic Cells ◽

Nk Cells ◽

Natural Killer ◽

Mhc Class Ii ◽

Nk Cell ◽

Phenotypic Differences ◽

Mhc Ii ◽

Class Ii Mhc

Interferon-producing killer dendritic cells (IKDCs) are a recently described subset of CD11cloB220+ cells that share phenotypic and functional properties of DCs and natural killer (NK) cells (Chan, C.W., E. Crafton, H.N. Fan, J. Flook, K. Yoshimura, M. Skarica, D. Brockstedt, T.W. Dubensky, M.F. Stins, L.L. Lanier, et al. 2006. Nat. Med. 12:207–213; Taieb, J., N. Chaput, C. Menard, L. Apetoh, E. Ullrich, M. Bonmort, M. Pequignot, N. Casares, M. Terme, C. Flament, et al. 2006. Nat. Med. 12:214–219). IKDC development appears unusual in that cytokines using the interleukin (IL)-2 receptor β (IL-2Rβ) chain but not those using the common γ chain (γc) are necessary for their generation. By directly comparing Rag2−/−γc−/y, Rag2−/−IL-2Rβ−/−, Rag2−/−IL-15−/−, and Rag2−/−IL-2−/− mice, we demonstrate that IKDC development parallels NK cell development in its strict IL-15 dependence. Moreover, IKDCs uniformly express NK-specific Ncr-1 transcripts (encoding NKp46), whereas NKp46+ cells are absent in Ncr1gfp/+γc−/y mice. Distinguishing features of IKDCs (CD11cloB220+MHC-II+) were carefully examined on developing NK cells in the bone marrow and on peripheral NK cells. As B220 expression was heterogeneous, defining B220lo versus B220hi NK1.1+ NK cells could be considered as arbitrary, and few phenotypic differences were noted between NK1.1+ NK cells bearing different levels of B220. CD11c expression did not correlate with B220 or major histocompatibility complex (MHC) class II (MHC-II) expression, and most MHC-II+ NK1.1+ cells did not express B220 and were thus not IKDCs. Finally, CD11c, MHC-II, and B220 levels were up-regulated on NK1.1+ cells upon activation in vitro or in vivo in a proliferation-dependent fashion. Our data suggest that the majority of CD11cloB220+ “IKDC-like” cells represent activated NK cells.

Download Full-text

African and Asian Zika Virus Isolates Display Phenotypic Differences Both In Vitro and In Vivo

American Journal of Tropical Medicine and Hygiene ◽

10.4269/ajtmh.17-0685 ◽

2018 ◽

Vol 98 (2) ◽

pp. 432-444 ◽

Cited By ~ 38

Author(s):

Darci R. Smith ◽

David A. Kulesh ◽

Stephanie A. Bellanca ◽

Andrew D. Haddow ◽

Thomas R. Sprague ◽

...

Keyword(s):

Zika Virus ◽

Phenotypic Differences ◽

Virus Isolates

Download Full-text

Replication Variance of African and Asian Lineage Zika Virus Strains in Different Cell Lines, Mosquitoes and Mice

Microorganisms ◽

10.3390/microorganisms9061250 ◽

2021 ◽

Vol 9 (6) ◽

pp. 1250

Author(s):

Tey Putita Ou ◽

Heidi Auerswald ◽

Saraden In ◽

Borin Peng ◽

Senglong Pang ◽

...

Keyword(s):

Zika Virus ◽

Mammalian Cells ◽

Vector Competence ◽

Mosquito Species ◽

Survival Rates ◽

Transmission Efficiency ◽

Phenotypic Differences ◽

Local Risk

Since the epidemic in 2007, studies on vector competence for Zika virus (ZIKV) have intensified, showing that the transmission efficiency varies depending on the vector population, ZIKV strain, and dose of the infectious blood meal. In this study, we aimed to investigate the replication of African and Asian ZIKV strains in vitro and in vivo in order to reveal their phenotypic differences. In addition, we investigated the vector competence of Cambodian Aedes aegypti (Ae. aegypti) mosquitoes (urban and rural) for these ZIKV strains. We observed a significantly higher pathogenicity of the African ZIKV strain in vitro (in mosquito and mammalian cells), and in vivo in both Ae. aegypti and mice. Both mosquito populations were competent to transmit ZIKV as early as 7 days p.i., depending on the population and the ZIKV strain. Ae. aegypti from rural habitats showed significant higher transmission and survival rates than those from urban. We observed the highest transmission efficiency for the African ZIKV isolate (93.3% 14 days p.i.) and for the Cambodian ZIKV isolate (80% 14 days p.i.). Overall, our results highlight the phenotypic differences of the ZIKV lineages and the potential risk of ZIKV transmission by Ae. aegypti mosquitoes. Further investigations of Cambodian mosquito species and ZIKV specific surveillance in humans is necessary in order to improve the local risk assessment.

Download Full-text