Portable on-chip colorimetric biosensing platform integrated with a smartphone for label/PCR-free detection of Cryptosporidium RNA

Cryptosporidium, a protozoan pathogen, is a leading threat to public health and the economy. Herein, we report the development of a portable, colorimetric biosensing platform for the sensitive, selective and label/PCR-free detection of Cryptosporidium RNA using oligonucleotides modified gold nanoparticles (AuNPs). A pair of specific thiolated oligonucleotides, complementary to adjacent sequences on Cryptosporidium RNA, were attached to AuNPs. The need for expensive laboratory-based equipment was eliminated by performing the colorimetric assay on a micro-fabricated chip in a 3D-printed holder assembly. A smartphone camera was used to capture an image of the color change for quantitative analysis. The detection was based on the aggregation of the gold nanoparticles due to the hybridization between the complementary Cryptosporidium RNA and the oligonucleotides immobilized on the AuNPs surface. In the complementary RNA’s presence, a distinctive color change of the AuNPs (from red to blue) was observed by the naked eye. However, in the presence of non-complementary RNA, no color change was observed. The sensing platform showed wide linear responses between 5 and 100 µM with a low detection limit of 5 µM of Cryptosporidium RNA. Additionally, the sensor developed here can provide information about different Cryptosporidium species present in water resources. This cost-effective, easy-to-use, portable and smartphone integrated on-chip colorimetric biosensor has great potential to be used for real-time and portable POC pathogen monitoring and molecular diagnostics.

Murine Irgm paralogs regulate non-redundant functions to execute host defense to Toxoplasma gondii

Gamma-interferon (IFNγ)-induced immunity-related GTPases (IRGs) confer cell-autonomous immunity to the intracellular protozoan pathogen Toxoplasma gondii . Effector IRGs are loaded onto the Toxoplasma -containing parasitophorous vacuole (PV), where they recruit ubiquitin ligases, ubiquitin-binding proteins, and IFNγ-inducible guanylate-binding proteins (Gbps), prompting PV lysis and parasite destruction. Host cells lacking the regulatory IRGs Irgm1 and Irgm3 fail to load effector IRGs, ubiquitin, and Gbps onto the PV and are consequently defective for cell-autonomous immunity to Toxoplasma . However, the role of the third regulatory IRG, Irgm2, in cell-autonomous immunity to Toxoplasma has remained unexplored. Here, we report that Irgm2 unexpectedly plays a limited role in the targeting of effector IRGs, ubiquitin, and Gbps to the Toxoplasma PV. Instead, Irgm2 is instrumental in the decoration of PVs with γ-aminobutyric acid receptor-associated protein-like 2 (GabarapL2). Cells lacking Irgm2 are as defective for cell-autonomous host defense to Toxoplasma as pan- Irgm -/- cells lacking all three Irgm proteins, and further, Irgm2 -/- mice succumb to Toxoplasma infections as readily as pan- Irgm -/- mice. These findings demonstrate that relative to Irgm1 and Irgm3, Irgm2 plays a distinct but critically important role in host resistance to Toxoplasma .

Nosema locustae (Protozoa, Microsporidia), a Biological Agent for Locust and Grasshopper Control

Effective locust and grasshopper control is crucial as locust invasions have seriously threatened crops and food security since ancient times. However, the preponderance of chemical insecticides, effective and widely used today, is increasingly criticized as a result of their adverse effects on human health and the environment. Alternative biological control methods are being actively sought to replace chemical pesticides. Nosema locustae (Synonyms: Paranosema locustae, Antonospora locustae), a protozoan pathogen of locusts and grasshoppers, was developed as a biological control agent as early as the 1980s. Subsequently, numerous studies have focused on its pathogenicity, host spectrum, mass production, epizootiology, applications, genomics, and molecular biology. Aspects of recent advances in N. locustae show that this entomopathogen plays a special role in locust and grasshopper management because it is safer, has a broad host spectrum of 144 orthopteran species, vertical transmission to offspring through eggs, long persistence in locust and grasshopper populations for more than 10 years, and is well adapted to various types of ecosystems in tropical and temperate regions. However, some limitations still need to be overcome for more efficient locust and grasshopper management in the future.

Pathologic and immunohistochemical findings in an outbreak of systemic toxoplasmosis in a mob of red kangaroos

Toxoplasma gondii is a zoonotic protozoan pathogen that infects many endothermic vertebrates, including humans; the domestic cat and other felids serve as the definitive host. Macropodids are considered highly susceptible to toxoplasmosis. Here, we describe the clinical, pathologic, and immunohistochemical findings of an outbreak of systemic toxoplasmosis in a mob of 11 red kangaroos ( Macropus rufus), with high morbidity (73%) and mortality (100%) rates. Affected animals had either severe and rapidly deteriorating clinical conditions or sudden death, which was correlated with widespread necrotizing lesions in multiple organs and intralesional T. gondii organisms identified via MIC3-specific immunohistochemistry and confirmed by REP529-specific rtPCR. Quantification of parasites demonstrated the highest parasite density in pulmonary parenchyma compared with other tissues. Our study highlights the continued importance of this severe condition in Australian marsupials.

Development of new dinuclear Fe(III) coordination compounds with in vitro nanomolar antitrypanosomal activity

Chagas disease is a neglected tropical disease caused by the protozoan pathogen Trypanosoma cruzi. The disease is the major public health problem affecting about 6 to 7 million people worldwide,...

Toxoplasma gondii GRA28 is required for specific induction of the regulatory chemokine CCL22 in human and mouse cells

ABSTRACTToxoplasma gondii is an intracellular protozoan pathogen of humans that causes severe disease in immunocompromised patients and in the developing fetus. T. gondii specifically alters production of the immunomodulatory chemokine CCL22 in human placental cells during infection. Using a combination of bioinformatics and molecular genetics, we have now identified T. gondii GRA28 as the gene product required for CCL22 induction. GRA28 is strongly co-regulated at the transcriptional level along with other known secreted effectors and their chaperones. GRA28 is secreted into the host cell where it localizes to the nucleus, and deletion of this gene results in reduced CCL22 secretion from human monocytes and second trimester placental explants. The impact of GRA28 on CCL22 is also conserved in mouse immune and placental cells and the deletion of GRA28 results in increased inflammatory responses and reduced CNS burden during mouse infectionsAUTHOR SUMMARYToxoplasma gondii is a globally ubiquitous pathogen that can cause severe disease in HIV/AIDS patients and can also cross the placenta and infect the developing fetus. We have found that placental and immune cells infected with T. gondii secrete signfiicant amounts of a chemokine (called “CCL22”) that is critical for immune tolerance during pregnancy. In order to better understand whether this is a response by the host or a process that is driven by the parasite, we have identified a T. gondii gene that is absolutely required to induce CCL22 production in human cells, indicating that CCL22 production is a process driven almost entirely by the parasite rather than the host. Consistent with its role in immune tolerance, we also found that T. gondii parasites lacking this gene are less able to proliferate and disseminate throughout the host. Taken together these data illustrate a direct relationship between CCL22 levels in the infected host and a key parasite effector, and provide an interesting example of how T. gondii can directly modulate host signaling pathways in order to facilitate its growth and dissemination.

First record of Crithidia expoeki (Trypanosomatida: Trypanosomatidae) from native Canadian bumble bees (Hymenoptera: Apidae: Bombus)

Bumble bees (Bombus Latrielle: Apidae) are important pollinators; however, declines of several species have been documented worldwide. Although pathogens have been linked to some declines, the biology, distribution, and impacts of most pathogens are poorly understood. Here, we report the first record of a recently characterized protozoan pathogen, Crithidia expoeki Schmid-Hempel & Tognazzo (Trypanosomatida: Trypanosomatidae), from bumble bees in Canada. This provides further insight on its global distribution and importance as a threat to bumble bees in Canada.