A Transfected Babesia bovis Parasite Line Expressing eGFP Is Able to Complete the Full Life Cycle of the Parasite in Mammalian and Tick Hosts

Johnson, Wendell C.; Hussein, Hala E.; Capelli-Peixoto, Janaína; Laughery, Jacob M.; Taus, Naomi S.; Suárez, Carlos E.; Ueti, Massaro W.

doi:10.3390/pathogens11060623

Cited by 4 publications

(3 citation statements)

References 31 publications

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“…At 30 days after Att-S74-T3Bo inoculation, vaccinated and control animals were challenged IV with one 1-ml stabilate containing 10 7 iRBC of the virulent homologous B. bovis strain Vir-S74-T3Bo. Previous studies have shown that a challenge dose of 10 7 iRBC of Vir-S74-T3Bo induces severe acute disease in calves with the onset of clinical signs starting approximately 9-10 days after infection ( 24 , 26 , 29 , 30 ). After challenge, all animals were monitored daily for clinical signs of acute bovine babesiosis, as described above.…”

Section: Methodsmentioning

confidence: 99%

Vaccination with an in vitro culture attenuated Babesia bovis strain safely protects highly susceptible adult cattle against acute bovine babesiosis

Bastos,

Capelli-Peixoto,

Laughery

et al. 2023

Front. Immunol.

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IntroductionLive in vivo attenuated Babesia bovis vaccines produced by sequential passages in splenectomized calves have historically been used to control acute bovine babesiosis in endemic areas worldwide. However, several constraints prevent the widespread use of these vaccines, including the need for several splenectomized calves to produce vaccine batches, and potential inconsistent parasite attenuation, which contraindicates their use for highly Babesia-susceptible adult cattle. Thus, the use of vaccines based on well-defined in vitro culture attenuated B. bovis strains emerges as a more sustainable and efficient alternative. Previous work demonstrated that the culture attenuated strain Att-S74-T3Bo is non-tick transmissible and able to safely protect calves against needle challenge with a B. bovis virulent strain.Methods and resultsHerein we evaluated safety and efficacy of Att-S74-T3Bo in preventing acute babesiosis in adult (>1.5 year of age) cattle. Results demonstrated that Att-S74-T3Bo vaccination of adult animals (n=5) induced self-limiting signs of acute infection and protected the vaccinated animals against challenge with the homologous virulent B. bovis strain Vir-S74-T3Bo. Att-S74-T3Bo-vaccinated adult cattle developed significant (P<0.05) monocytosis, with concomitant neutropenia and CD4+ leukopenia, in peripheral blood early after vaccination. Also, vaccinated animals developed a specific signature of pro- and anti-inflammatory cytokine expression in peripheral blood and significant levels of IgM, total IgG, IgG1, and IgG2 against the B. bovis immunodominant antigen RAP-1 CT. Strikingly, none of the vaccinated animals showed any signs of acute babesiosis after challenge with Vir-S74-T3Bo. In contrast, control adult cattle (n=5) showed pathognomonic symptoms of acute babesiosis, and significant decrease (P<0.05) in lymphocytes, monocytes, and neutrophils, starting on day 7 post-challenge. All control animals developed severe acute disease and were euthanized on days 10 through 12 days post-challenge.Discussion and conclusionEvidence from this study indicates that Att-S74-T3Bo safely protects highly susceptible adult cattle against challenge with a homologous virulent strain of B. bovis. In conclusion, Att-S74-T3Bo may be considered as a potential efficient and sustainable attenuated candidate vaccine strain to control acute bovine babesiosis in highly susceptible adult cattle. Future studies should focus on increasing the number of animals vaccinated, duration of immunity, and efficacy of this attenuated strain against heterologous virulent parasite strains.

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Section: Methodsmentioning

confidence: 99%

Vaccination with an in vitro culture attenuated Babesia bovis strain safely protects highly susceptible adult cattle against acute bovine babesiosis

Bastos,

Capelli-Peixoto,

Laughery

et al. 2023

Front. Immunol.

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“…Following the genome sequencing of the B. bovis T2Bo strain (26), a marked increase in understanding the parasite's biology has emerged from functional studies using genetically modified parasites and high-throughput approaches (27)(28)(29)(30)(31)(32). Therefore, several novel parasite blood and tick stage antigens have been discovered and evaluated in vaccine trials (31,(33)(34)(35).…”

Section: Introductionmentioning

confidence: 99%

Identification of novel immune correlates of protection against acute bovine babesiosis by superinfecting cattle with in vitro culture attenuated and virulent Babesia bovis strains

et al. 2022

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The apicomplexan tickborne parasites Babesia bovis and B. bigemina are the major causative agents of bovine babesiosis, a disease that negatively affects the cattle industry and food safety around the world. The absence of correlates of protection represents one major impediment for the development of effective and sustainable vaccines against bovine babesiosis. Herein we superinfected cattle with attenuated and virulent strains of B. bovis to investigate immune correlates of protection against acute bovine babesiosis. Three 6-month-old Holstein calves were infected intravenously (IV) with the in vitro culture attenuated Att-S74-T3Bo B. bovis strain (106 infected bovine red blood cells (iRBC)/calf) while three age-matched Holstein calves were inoculated IV with normal RBC as controls (106 RBC/calf). All Att-S74-T3Bo-infected calves showed a significant increase in temperature early after inoculation but recovered without treatment. Att-S74-T3Bo-infected calves also developed: (a) monocytosis, neutropenia, and CD4+ lymphopenia in peripheral blood on days 3 to 7 post-inoculation; (b) significant levels of TNFα, CXCL10, IFNγ, IL-4, and IL-10 in sera at day 6 after infection; and (c) IgM and IgG against B. bovis antigens, starting at days 10 and 30 post-inoculation, respectively. At 46 days post-Att-S74-T3Bo inoculation, all experimental calves were infected IV with the homologous virulent B. bovis strain Vir-S74-T3Bo (107 iRBC/calf). All Att-S74-T3Bo-infected calves survived superinfection with Vir-S74-T3Bo without displaying signs of acute babesiosis. In contrast, control animals showed signs of acute disease, starting at day 10 post-Vir-S74-T3Bo infection, and two of them were humanely euthanized at days 13 and 14 after inoculation due to the severity of their symptoms. Also, control calves showed higher (P<0.05) parasite load in peripheral blood compared to animals previously exposed to Att-S74-T3Bo. No significant alterations in the profile of leukocytes and cytokines were observed in Att-S74-T3Bo-inoculated after Vir-S74-T3Bo infection. In conclusion, data demonstrate novel changes in the profile of blood immune cells and cytokine expression in peripheral blood that are associated with protection against acute bovine babesiosis. These identified immune correlates of protection may be useful for designing effective and sustainable vaccines against babesiosis in cattle.

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“…A transfected B. bovis line tfBbo5480 expressing eGFP [ 23 ] was used to generate sexual stages for immunofluorescence assay (IFA). Live B. bovis male and female-enriched gametes separated from induced sexual stage cultures were washed in PBS plus 3% bovine serum albumin (BSA).…”

Section: Methodsmentioning

confidence: 99%

Expression of sex-specific molecular markers by Babesia bovis gametes

Hussein,

Johnson,

Taus

et al. 2024

Parasites Vectors

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Background Bovine babesiosis caused by Babesia bovis is one of the most important tick-borne diseases of cattle in tropical and subtropical regions. Babesia bovis parasites have a complex lifecycle, including development within the mammalian host and tick vector. In the tick midgut, extracellular Babesia parasites transform into gametes that fuse to form zygotes. To date, little is known about genes and proteins expressed by male gametes. Methods and results We developed a method to separate male gametes from in vitro induced B. bovis culture. Separation enabled the validation of sex-specific markers. Collected male gametocytes were observed by Giemsa-stained smear and live-cell fluorescence microscopy. Babesia male gametes were used to confirm sex-specific markers by quantitative real-time PCR. Some genes were found to be male gamete specific genes including pka, hap2, α-tubulin II and znfp2. However, α-tubulin I and ABC transporter, trap2-4 and ccp1-3 genes were found to be upregulated in culture depleted of male gametes (female-enriched). Live immunofluorescence analysis using polyclonal antibodies confirmed surface expression of HAP2 by male and TRAP2-4 by female gametes. These results revealed strong markers to distinguish between B. bovis male and female gametes. Conclusions Herein, we describe the identification of sex-specific molecular markers essential for B. bovis sexual reproduction. These tools will enhance our understanding of the biology of sexual stages and, consequently, the development of additional strategies to control bovine babesiosis. Graphical Abstract

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A Transfected Babesia bovis Parasite Line Expressing eGFP Is Able to Complete the Full Life Cycle of the Parasite in Mammalian and Tick Hosts

Cited by 4 publications

References 31 publications

Vaccination with an in vitro culture attenuated Babesia bovis strain safely protects highly susceptible adult cattle against acute bovine babesiosis

Vaccination with an in vitro culture attenuated Babesia bovis strain safely protects highly susceptible adult cattle against acute bovine babesiosis

Identification of novel immune correlates of protection against acute bovine babesiosis by superinfecting cattle with in vitro culture attenuated and virulent Babesia bovis strains

Expression of sex-specific molecular markers by Babesia bovis gametes

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