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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV120157	1/1	Bartonella henselae	Bacteria	(d)(U)C Filtration	Roden JA	2012	29%
Study summary Full title Hemin binding protein C is found in outer membrane vesicles and protects Bartonella henselae against toxic concentrations of hemin All authors Roden JA, Wells DH, Chomel BB, Kasten RW, Koehler JE Journal Infect Immun Abstract Bartonella species are gram-negative, emerging bacterial pathogens found in two distinct environment (show more...)Bartonella species are gram-negative, emerging bacterial pathogens found in two distinct environments. In the gut of the obligately hematophagous arthropod vector, bartonellae are exposed to concentrations of heme that are toxic to other bacteria. In the bloodstream of the mammalian host, access to heme and iron is severely restricted. Bartonellae have unusually high requirements for heme, which is their only utilizable source of iron. Although heme is essential for Bartonella survival, little is known about genes involved in heme acquisition and detoxification. We developed a strategy for high-efficiency transposon mutagenesis to screen for genes in B. henselae heme binding and uptake pathways. We identified a B. henselae transposon mutant that constitutively expresses the hemin binding protein C (hbpC) gene. In the wild-type strain, transcription of B. henselae hbpC was upregulated at arthropod temperature (28°C), compared to mammalian temperature (37°C). In the mutant strain, temperature-dependent regulation was absent. We demonstrated that HbpC binds hemin and localizes to the B. henselae outer membrane and outer membrane vesicles. Overexpression of hbpC in B. henselae increased resistance to heme toxicity, implicating HbpC in protection of B. henselae from the toxic levels of heme present in the gut of the arthropod vector. Experimental inoculation of cats with B. henselae strains demonstrated that both constitutive expression and deletion of hbpC affect the ability of B. henselae to infect the cat host. Modulation of hbpC expression appears to be a strategy employed by B. henselae to survive in the arthropod vector and the mammalian host. (hide) EV-METRIC 29% (76th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Bacteria Sample origin NAY Focus vesicles OMV Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Filtration Adj. k-factor 139.8 (pelleting) Protein markers EV: non-EV: Proteomics yes Show all info Study aim Other/Function of a bacterial protein in infection Sample Species Bartonella henselae Sample Type Bacteria Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: rotor type 45Ti Pelleting: adjusted k-factor 139.8 Filtration steps 0.22µm or 0.2µm Characterization: Protein analysis Characterization: Particle analysis EM EM-type transmission EM Image type Wide-field

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EV-TRACK ID	EV120157
species	Bartonella henselae
sample type	Bacteria
condition	NAY
separation protocol	(d)(U)C Filtration
Exp. nr.	1
EV-METRIC %	29