Vibrational Spectroscopic Based Approach for Diagnosing Babesia bovis Infection.

Babesia bovis parasites present a serious and significant health concern for the beef and dairy industries in many parts of the world. Difficulties associated with the current diagnostic techniques include the following: they are prone to human error (microscopy) or expensive and time-consuming (polymerase chain reaction) to perform. Little is known about the biochemical changes in blood that are associated with Babesia infections. The discovery of new biomarkers will lead to improved diagnostic outcomes for the cattle industry. Vibrational spectroscopic technologies can record a chemical snapshot of the entire organism and the surrounding cell thereby providing a phenotype of the organism and the host infected cell. Here, we demonstrate the applicability of vibrational spectroscopic imaging techniques including Atomic Force Microscopy Infrared (AFM-IR) and confocal Raman microscopy to discover new biomarkers for B. bovis infections. Furthermore, we applied Attenuated Total Reflection Fourier Transform Infrared (ATR-FTIR) to detect B. bovis in red blood cells (RBCs). Based on changes in the IR spectral bands, with ATR-FTIR in combination with Partial Least Squares-Discriminant Analysis we were able to discriminate infected samples from controls with a sensitivity and specificity of 92.0% and 91.7%, respectively, in less than 2 min, excluding sample extraction and preparation. The proposed method utilized a lysis approach to remove hemoglobin from the suspension of infected and uninfected cells, which significantly increased the sensitivity and specificity compared to measurements performed on intact infected red blood cells (intact infected RBC, 77.3% and 79.2%). This work represents a holistic spectroscopic study from the level of the single infected RBC using AFM-IR and confocal Raman to the detection of the parasite in a cell population using ATR-FTIR for a babesiosis diagnostic.

Babesiosis Vaccines: Lessons Learned, Challenges Ahead, and Future Glimpses.

The incidence and prevalence of babesiosis in animals and humans is increasing, yet prevention, control, or treatment measures remain limited and ineffective. Despite a growing body of new knowledge of the biology, pathogenicity, and virulence of Babesia parasites, there is still no well-defined, adequately effective and easily deployable vaccine. While numerous published studies suggest that the development of such anti-Babesia vaccines should be feasible, many others identify significant challenges that need to be overcome in order to succeed. Here, we review historic and recent attempts in babesiosis vaccine discovery to avoid past pitfalls, learn new lessons, and provide a roadmap to guide the development of next-generation babesiosis vaccines.

Unravelling the cellular and molecular pathogenesis of bovine babesiosis: is the sky the limit?

The global impact of bovine babesiosis caused by the tick-borne apicomplexan parasites Babesia bovis, Babesia bigemina and Babesia divergens is vastly underappreciated. These parasites invade and multiply asexually in bovine red blood cells (RBCs), undergo sexual reproduction in their tick vectors (Rhipicephalus spp. for B. bovis and B. bigemina, and Ixodes ricinus for B. divergens) and have a trans-ovarial mode of transmission. Babesia parasites can cause acute and persistent infections to adult naïve cattle that can occur without evident clinical signs, but infections caused by B. bovis are associated with more severe disease and increased mortality, and are considered to be the most virulent agent of bovine babesiosis. In addition, babesiosis caused by B. divergens has an important zoonotic potential. The disease caused by B. bovis and B. bigemina can be controlled, at least in part, using therapeutic agents or vaccines comprising live-attenuated parasites, but these methods are limited in terms of their safety, ease of deployability and long-term efficacy, and improved control measures are urgently needed. In addition, expansion of tick habitats due to climate change and other rapidly changing environmental factors complicate efficient control of these parasites. While the ability to cause persistent infections facilitates transmission and persistence of the parasite in endemic regions, it also highlights their capacity to evade the host immune responses. Currently, the mechanisms of immune responses used by infected bovines to survive acute and chronic infections remain poorly understood, warranting further research. Similarly, molecular details on the processes leading to sexual reproduction and the development of tick-stage parasites are lacking, and such tick-specific molecules can be targets for control using alternative transmission blocking vaccines. In this review, we identify and examine key phases in the life-cycle of Babesia parasites, including dependence on a tick vector for transmission, sexual reproduction of the parasite in the midgut of the tick, parasite-dependent invasion and egression of bovine RBCs, the role of the spleen in the clearance of infected RBCs (IRBCs), and age-related disease resistance in cattle, as opportunities for developing improved control measures. The availability of integrated novel research approaches including "omics" (such as genomics, transcriptomics, and proteomics), gene modification, cytoadhesion assays, RBC invasion assays and methods for in vitro induction of sexual-stage parasites will accelerate our understanding of parasite vulnerabilities. Further, producing new knowledge on these vulnerabilities, as well as taking full advantage of existing knowledge, by filling important research gaps should result in the development of next-generation vaccines to control acute disease and parasite transmission. Creative and effective use of current and future technical and computational resources are needed, in the face of the numerous challenges imposed by these highly evolved parasites, for improving the control of this disease. Overall, bovine babesiosis is recognised as a global disease that imposes a serious burden on livestock production and human livelihood, but it largely remains a poorly controlled disease in many areas of the world. Recently, important progress has been made in our understanding of the basic biology and host-parasite interactions of Babesia parasites, yet a good deal of basic and translational research is still needed to achieve effective control of this important disease and to improve animal and human health.

Evaluation of 4-Amino 2-Anilinoquinazolines against Plasmodium and Other Apicomplexan Parasites In Vitro and in a P. falciparum Humanized NOD-scid IL2Rγnull Mouse Model of Malaria.

A series of 4-amino 2-anilinoquinazolines optimized for activity against the most lethal malaria parasite of humans, Plasmodium falciparum, was evaluated for activity against other human Plasmodium parasites and related apicomplexans that infect humans and animals. Four of the most promising compounds from the 4-amino 2-anilinoquinazoline series were equally as effective against the asexual blood stages of the zoonotic P. knowlesi, suggesting that they could also be effective against the closely related P. vivax, another important human pathogen. The 2-anilinoquinazoline compounds were also potent against an array of P. falciparum parasites resistant to clinically available antimalarial compounds, although slightly less so than against the drug-sensitive 3D7 parasite line. The apicomplexan parasites Toxoplasma gondii, Babesia bovis, and Cryptosporidium parvum were less sensitive to the 2-anilinoquinazoline series with a 50% effective concentration generally in the low micromolar range, suggesting that the yet to be discovered target of these compounds is absent or highly divergent in non-Plasmodium parasites. The 2-anilinoquinazoline compounds act as rapidly as chloroquine in vitro and when tested in rodents displayed a half-life that contributed to the compound's capacity to clear P. falciparum blood stages in a humanized mouse model. At a dose of 50 mg/kg of body weight, adverse effects to the humanized mice were noted, and evaluation against a panel of experimental high-risk off targets indicated some potential off-target activity. Further optimization of the 2-anilinoquinazoline antimalarial class will concentrate on improving in vivo efficacy and addressing adverse risk.