scholarly journals Identification of a Novel Elastin-Degrading Enzyme from the Fish PathogenFlavobacterium psychrophilum

2019 ◽  
Vol 85 (6) ◽  
Author(s):  
T. Rochat ◽  
D. Pérez-Pascual ◽  
H. Nilsen ◽  
M. Carpentier ◽  
S. Bridel ◽  
...  
Keyword(s):  
Cell Surface ◽  
Pathogenic Bacteria ◽  
Bacterial Pathogen ◽  
Salmonid Fish ◽  
Economic Losses ◽  
Gene Encoding ◽  
Content Type ◽  
Flavobacterium Psychrophilum ◽  
A Genome ◽  
A Cell

ABSTRACTHydrolytic extracellular enzymes degrading host tissues potentially play a role in bacterial pathogenesis.Flavobacterium psychrophilumis an important bacterial pathogen of salmonid fish reared in freshwater throughout the world. Diversity among isolates has been described at the phenotypic, serological, and genomic levels, but the links between these various traits remain poorly understood. Using a genome-wide association study, we identified a gene encoding a novel elastinolytic enzyme inF. psychrophilum. To formally demonstrate enzymatic activity, this gene (FP0506from strain JIP 02/86) was expressed in the elastinolysis-deficient strain OSU THCO2-90, resulting in proficient elastin-degrading cells. The encoded protein is predicted to be a cell-surface-exposed lipoprotein with no homology to previously reported elastases. FP0506 might belong to the zincin tribe and gluzincin clan of metalloproteases, and this new elastase-encoding gene seems to be present only in some members of the familyFlavobacteriaceae.IMPORTANCEElastin is an important proteinaceous component of vertebrate connective tissues (e.g., blood vessels, lung, and skin), to which it confers elasticity. Elastases have been identified in a number of pathogenic bacteria. They are thought to be required for tissue penetration and dissemination, acting as “spreading factors.”Flavobacterium psychrophilumis a devastating bacterial pathogen of salmonid fish (salmon and trout) that is responsible for severe economic losses worldwide. This pathogen displays strong proteolytic activities. Using a variety of techniques, including genome comparisons, we identified a gene encoding a novel elastase inF. psychrophilum. The encoded protein is predicted to be a cell-surface-exposed lipoprotein with no homology to previously reported elastases. In addition, this elastase likely belongs to a new family of proteases that seems to be present only in some members of this important group of bacteria.

Cloning of the gene encoding TROP-2, a cell-surface glycoprotein expressed by human carcinomas

1995 ◽  
Vol 62 (5) ◽  
pp. 610-618 ◽  
Author(s):  
Mara Fornaro ◽  
Roberta Dell' Arciprete ◽  
Manuela Stella ◽  
Cecilia Bucci ◽  
Michele Nutini ◽  
...  
Keyword(s):  
Cell Surface ◽  
Surface Glycoprotein ◽  
Gene Encoding ◽  
Cell Surface Glycoprotein ◽  
Human Carcinomas ◽  
A Cell

scholarly journals Modulating Pathogenesis with Mobile-CRISPRi

2019 ◽  
Vol 201 (22) ◽  
Author(s):  
Jiuxin Qu ◽  
Neha K. Prasad ◽  
Michelle A. Yu ◽  
Shuyan Chen ◽  
Amy Lyden ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pathogenic Bacteria ◽  
Bacterial Species ◽  
Effector Proteins ◽  
Infection Model ◽  
Secretion Systems ◽  
Bacterial Genomes ◽  
Gene Encoding ◽  
Content Type ◽  
Animal Infection

ABSTRACT Conditionally essential (CE) genes are required by pathogenic bacteria to establish and maintain infections. CE genes encode virulence factors, such as secretion systems and effector proteins, as well as biosynthetic enzymes that produce metabolites not found in the host environment. Due to their outsized importance in pathogenesis, CE gene products are attractive targets for the next generation of antimicrobials. However, the precise manipulation of CE gene expression in the context of infection is technically challenging, limiting our ability to understand the roles of CE genes in pathogenesis and accordingly design effective inhibitors. We previously developed a suite of CRISPR interference-based gene knockdown tools that are transferred by conjugation and stably integrate into bacterial genomes that we call Mobile-CRISPRi. Here, we show the efficacy of Mobile-CRISPRi in controlling CE gene expression in an animal infection model. We optimize Mobile-CRISPRi in Pseudomonas aeruginosa for use in a murine model of pneumonia by tuning the expression of CRISPRi components to avoid nonspecific toxicity. As a proof of principle, we demonstrate that knock down of a CE gene encoding the type III secretion system (T3SS) activator ExsA blocks effector protein secretion in culture and attenuates virulence in mice. We anticipate that Mobile-CRISPRi will be a valuable tool to probe the function of CE genes across many bacterial species and pathogenesis models. IMPORTANCE Antibiotic resistance is a growing threat to global health. To optimize the use of our existing antibiotics and identify new targets for future inhibitors, understanding the fundamental drivers of bacterial growth in the context of the host immune response is paramount. Historically, these genetic drivers have been difficult to manipulate precisely, as they are requisite for pathogen survival. Here, we provide the first application of Mobile-CRISPRi to study conditionally essential virulence genes in mouse models of lung infection through partial gene perturbation. We envision the use of Mobile-CRISPRi in future pathogenesis models and antibiotic target discovery efforts.

scholarly journals The Acinetobacter baumannii Znu System Overcomes Host-Imposed Nutrient Zinc Limitation

2019 ◽  
Vol 87 (12) ◽  
Author(s):  
Laura E. Hesse ◽  
Zachery R. Lonergan ◽  
William N. Beavers ◽  
Eric P. Skaar
Keyword(s):  
Acinetobacter Baumannii ◽  
Defense Mechanisms ◽  
Pathogenic Bacteria ◽  
Bacterial Pathogen ◽  
Multidrug Resistant ◽  
World Health ◽  
Content Type ◽  
Nutritional Immunity ◽  
Essential Nutrient ◽  
Health Organization

ABSTRACT Acinetobacter baumannii is an opportunistic bacterial pathogen capable of causing a variety of infections, including pneumonia, sepsis, wound, and burn infections. A. baumannii is an increasing threat to public health due to the prevalence of multidrug-resistant strains, leading the World Health Organization to declare A. baumannii a “Priority 1: Critical” pathogen, for which the development of novel antimicrobials is desperately needed. Zinc (Zn) is an essential nutrient that pathogenic bacteria, including A. baumannii, must acquire from their hosts in order to survive. Consequently, vertebrate hosts have defense mechanisms to sequester Zn from invading bacteria through a process known as nutritional immunity. Here, we describe a Zn uptake (Znu) system that enables A. baumannii to overcome this host-imposed Zn limitation. The Znu system consists of an inner membrane ABC transporter and an outer membrane TonB-dependent receptor. Strains of A. baumannii lacking any individual Znu component are unable to grow in Zn-starved conditions, including in the presence of the host nutritional immunity protein calprotectin. The Znu system contributes to Zn-limited growth by aiding directly in the uptake of Zn into A. baumannii cells and is important for pathogenesis in murine models of A. baumannii infection. These results demonstrate that the Znu system allows A. baumannii to subvert host nutritional immunity and acquire Zn during infection.

scholarly journals Identification of Strain-Specific Sequences That Distinguish a Mycoplasma gallisepticum Vaccine Strain from Field Isolates

2016 ◽  
Vol 55 (1) ◽  
pp. 244-252 ◽  
Author(s):  
Camir Ricketts ◽  
Larissa Pickler ◽  
John Maurer ◽  
Saravanaraj Ayyampalayam ◽  
Maricarmen García ◽  
...  
Keyword(s):  
Vaccine Strain ◽  
Reference Genome ◽  
Hypothetical Protein ◽  
Mycoplasma Gallisepticum ◽  
Economic Losses ◽  
Poultry Production ◽  
Content Type ◽  
Field Isolates ◽  
Avian Mycoplasmosis ◽  
A Cell

ABSTRACTDespite attempts to control avian mycoplasmosis through management, vaccination, and surveillance,Mycoplasma gallisepticumcontinues to cause significant morbidity, mortality, and economic losses in poultry production. Live attenuated vaccines are commonly used in the poultry industry to control avian mycoplasmosis; unfortunately, some vaccines may revert to virulence and vaccine strains are generally difficult to distinguish from natural field isolates. In order to identify genome differences among vaccine revertants, vaccine strains, and field isolates, whole-genome sequencing of theM. gallisepticumvaccine strain ts-11 and several “ts-11-like” strains isolated from commercial flocks was performed using Illumina and 454 pyrosequencing and the sequenced genomes compared to theM. gallisepticumRlowreference genome. The collective contigs for each strain were annotated using the fully annotatedMycoplasmareference genome. The analysis revealed genetic differences amongvlhAalleles, as well as among genes annotated as coding for a cell wall surface anchor protein (mg0377) and a hypothetical protein gene,mg0359, unique toM. gallisepticumts-11 vaccine strain. PCR protocols were designed to target 5 sequences unique to theM. gallisepticumts-11 strain:vlhA3.04a,vlhA3.04b,vlhA3.05,mg0377, andmg0359. All ts-11 isolates were positive for the five gene alleles tested by PCR; however, 5 to 36% of field isolates were also positive for at least one of the alleles tested. A combination of PCR tests forvlhA3.04a,vlhA3.05, andmg0359was able to distinguish theM. gallisepticumts-11 vaccine strain from field isolates. This method will further supplement current approaches to quickly distinguishM. gallisepticumvaccine strains from field isolates.

scholarly journals Brussowvirus SW13 Requires a Cell Surface-Associated Polysaccharide To Recognize Its Streptococcus thermophilus Host

2022 ◽  
Vol 88 (1) ◽  
Author(s):  
Katherine Lavelle ◽  
Irina Sadovskaya ◽  
Evgeny Vinogradov ◽  
Philip Kelleher ◽  
Gabriele A. Lugli ◽  
...  
Keyword(s):  
Cell Surface ◽  
Product Quality ◽  
Starter Culture ◽  
Streptococcus Thermophilus ◽  
Content Type ◽  
Fermentation Processes ◽  
A Cell ◽  
Dairy Fermentation

Streptococcus thermophilus is an important starter culture bacterium in global dairy fermentation processes, where it is used for the production of various cheeses and yogurt. Bacteriophage predation of the species can result in substandard product quality and, in rare cases, complete fermentation collapse.

scholarly journals Complete Genome Sequence of Acinetobacter baumannii Phage BS46

2020 ◽  
Vol 9 (22) ◽  
Author(s):  
Anastasia V. Popova ◽  
Mikhail M. Shneider ◽  
Yulia V. Mikhailova ◽  
Andrey A. Shelenkov ◽  
Dmitry A. Shagin ◽  
...  
Keyword(s):  
Complete Genome Sequence ◽  
Complete Genome ◽  
Open Reading Frames ◽  
Capsular Polysaccharides ◽  
Bacterial Host ◽  
Gene Encoding ◽  
Content Type ◽  
Double Stranded Dna ◽  
A Genome ◽  
Reading Frames

ABSTRACT Acinetobacter myovirus BS46 was isolated from sewage by J. S. Soothill in 1991. We have sequenced the genome of BS46 and found it to be almost unique. BS46 contains double-stranded DNA with a genome size of 94,068 bp and 176 predicted open reading frames. The gene encoding the tailspike that presumably possesses depolymerase activity toward the capsular polysaccharides of the bacterial host was identified.

scholarly journals Large-Scale Analysis ofFlavobacterium psychrophilumMultilocus Sequence Typing Genotypes Recovered from North American Salmonids Indicates that both Newly Identified and Recurrent Clonal Complexes Are Associated with Disease

2019 ◽  
Vol 85 (6) ◽  
Author(s):  
Christopher Knupp ◽  
Gregory D. Wiens ◽  
Mohamed Faisal ◽  
Douglas R. Call ◽  
Kenneth D. Cain ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Rainbow Trout ◽  
North America ◽  
North American ◽  
Economic Losses ◽  
Content Type ◽  
Flavobacterium Psychrophilum ◽  
Coldwater Disease ◽  
Rainbow Trout Fry Syndrome ◽  
Bacterial Coldwater Disease

ABSTRACTFlavobacterium psychrophilum, the etiological agent of bacterial coldwater disease (BCWD) and rainbow trout fry syndrome (RTFS), causes significant economic losses in salmonid aquaculture, particularly in rainbow trout (Oncorhynchus mykiss). Prior studies have used multilocus sequence typing (MLST) to examine genetic heterogeneity withinF. psychrophilum. At present, however, its population structure in North America is incompletely understood, as only 107 isolates have been genotyped. Herein, MLST was used to investigate the genetic diversity of an additional 314 North AmericanF. psychrophilumisolates that were recovered from ten fish host species from 20 U.S. states and 1 Canadian province over nearly four decades. These isolates were placed into 66 sequence types (STs), 47 of which were novel, increasing the number of clonal complexes (CCs) in North America from 7 to 12. Newly identified CCs were diverse in terms of host association, distribution, and association with disease. The largestF. psychrophilumCC identified was CC-ST10, within which 10 novel genotypes were discovered, most of which came fromO. mykissexperiencing BCWD. This discovery, among others, provides evidence for the hypothesis that ST10 (i.e., the founding ST of CC-ST10) originated in North America. Furthermore, ST275 (in CC-ST10) was recovered from wild/feral adult steelhead and marks the first recovery of CC-ST10 from wild/feral fish in North America. Analyses also revealed that at the allele level, the diversification ofF. psychrophilumin North America is driven three times more frequently by recombination than random nucleic acid mutation, possibly indicating how new phenotypes emerge within this species.IMPORTANCEFlavobacterium psychrophilumis the causative agent of bacterial coldwater disease (BCWD) and rainbow trout fry syndrome (RTFS), both of which cause substantial losses in farmed fish populations worldwide. To better prevent and control BCWD and RTFS outbreaks, we sought to characterize the genetic diversity of several hundredF. psychrophilumisolates that were recovered from diseased fish across North America. Results highlighted multipleF. psychrophilumgenetic strains that appear to play an important role in disease events in North American aquaculture facilities and suggest that the practice of trading fish eggs has led to the continental and transcontinental spread of this bacterium. The knowledge generated herein will be invaluable toward guiding the development of future disease prevention techniques.

scholarly journals High Genetic Diversity in Flavobacterium psychrophilum Isolates from Healthy Rainbow Trout (Oncorhynchus mykiss) Farmed in the Same Watershed, Revealed by Two Typing Methods

2020 ◽  
Vol 87 (2) ◽  
Author(s):  
Ségolène Calvez ◽  
Nora Navarro-Gonzalez ◽  
Charlène Siekoula-Nguedia ◽  
Catherine Fournel ◽  
Eric Duchaud
Keyword(s):  
Genetic Diversity ◽  
Rainbow Trout ◽  
Oncorhynchus Mykiss ◽  
Clonal Complex ◽  
Economic Losses ◽  
High Genetic Diversity ◽  
Content Type ◽  
Flavobacterium Psychrophilum ◽  
Sequence Types ◽  
Apparently Healthy

ABSTRACT Flavobacterium psychrophilum affects salmonid health worldwide and causes economic losses. The genetic diversity of the pathogen must be considered to develop control methods. However, previous studies have reported both high and low levels of genetic diversity. The present longitudinal study aimed at assessing the genetic diversity of F. psychrophilum at a small temporal and geographic scale. Four farms located on the same watershed in France were studied. Rainbow trout (Oncorhynchus mykiss) batches were monitored, and apparently healthy individuals were sampled over 1 year. A total of 288 isolates were recovered from fish organs (gills and spleen) and eggs. Pulsed field gel electrophoresis revealed high genetic diversity. Multilocus sequence typing performed on a selection of 31 isolates provided congruent results, as follows: 18 sequence types (STs) were found, of which 13 were novel. The mean gene diversity (H = 0.8413) was much higher than that previously reported for this host species, although the sampling was restricted to a single watershed and 1 year. Seven isolates out of 31 were assigned to clonal complex ST10 (CC-ST10), which is the predominant clonal complex in the main salmonid production areas. A split decomposition tree reflected a panmictic population. This finding is important for aquaculture veterinarians in their diagnostic procedure, as the choice of adequate antibiotic treatment is conditioned by the correct identification of the causative agent. Furthermore, this study expands our knowledge on genetic diversity required for the development of an effective vaccine against F. psychrophilum. IMPORTANCE The bacterium Flavobacterium psychrophilum is a serious pathogen in many fish species, especially salmonids, that is responsible for considerable economic losses worldwide. In order to treat infections and to develop vaccines, the genetic diversity of this bacterium needs to be known. We assessed the genetic diversity of F. psychrophilum isolates from apparently healthy rainbow trout raised in several fish farms in the same watershed in France. Two different genotyping methods revealed high diversity. The majority of isolates were unrelated to clonal complex sequence type 10 (CC-ST10), the clonal complex that is predominant worldwide and associated with disease in rainbow trout. In addition, we found 13 novel sequence types. These results suggest that a diverse subpopulation of F. psychrophilum may be harbored by rainbow trout.

scholarly journals Gene Deletion Strategy To Examine the Involvement of the Two Chondroitin Lyases in Flavobacterium columnare Virulence

2015 ◽  
Vol 81 (21) ◽  
pp. 7394-7402 ◽  
Author(s):  
Nan Li ◽  
Ting Qin ◽  
Xiao Lin Zhang ◽  
Bei Huang ◽  
Zhi Xin Liu ◽  
...  
Keyword(s):  
Virulence Factors ◽  
Gene Deletion ◽  
Genetic Manipulation ◽  
Bacterial Pathogen ◽  
Infected Fish ◽  
Economic Losses ◽  
Flavobacterium Columnare ◽  
Wild Type ◽  
Single Strain ◽  
Content Type

ABSTRACTFlavobacterium columnareis an important bacterial pathogen of freshwater fish that causes high mortality of infected fish and heavy economic losses in aquaculture. The pathogenesis of this bacterium is poorly understood, in part due to the lack of efficient methods for genetic manipulation. In this study, a gene deletion strategy was developed and used to determine the relationship between the production of chondroitin lyases and virulence. TheF. johnsoniaeompApromoter (PompA) was fused tosacBto construct a counterselectable marker forF. columnare.F. columnarecarrying PompA-sacBfailed to grow on media containing 10% sucrose. A suicide vector carrying PompA-sacBwas constructed, and a gene deletion strategy was developed. Using this approach, the chondroitin lyase-encoding genes,cslAandcslB, were deleted. The ΔcslAand ΔcslBmutants were both partially deficient in digestion of chondroitin sulfate A, whereas a double mutant (ΔcslAΔcslB) was completely deficient in chondroitin lyase activity. Cells ofF. columnarewild-type strain G4and of the chondroitin lyase-deficient ΔcslAΔcslBmutant exhibited similar levels of virulence toward grass carp in single-strain infections. Coinfections, however, revealed a competitive advantage for the wild type over the chondroitin lyase mutant. The results indicate that chondroitin lyases are not essential virulence factors ofF. columnarebut may contribute to the ability of the pathogen to compete and cause disease in natural infections. The gene deletion method developed in this study may be employed to investigate the virulence factors of this bacterium and may have wide application in many other members of the phylumBacteroidetes.

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