Pathogen- and host-directed pharmacologic strategies for control of Vairimorpha (Nosema) spp. infection in honey bees.

Microsporidia are obligate intracellular parasites of the Fungal Kingdom that cause widespread infections in nature, with important effects on invertebrates involved in food production systems. The two microsporidian species Vairimorpha (Nosema) ceranae (and the less common Vairimorpha (Nosema) apis) can cause individual disease in honey bees and contribute to colony collapse. The efficacy, safety, and availability of fumagillin, the only drug currently approved to treat microsporidia infection in bees, is uncertain. In this review, we will discuss some of the most promising alternative strategies for the mitigation of Vairimorpha spp. with an emphasis on infection by V. ceranae, now the dominant species infecting bees. We will focus on pharmacologic interventions where the mechanism of action is known and examine both pathogen-directed and host-directed approaches. As limiting toxicity to host cells has been especially emphasized in treating bees that are already facing numerous stressors, strategies that disrupt pathogen-specific targets may be especially advantageous. Therefore, efforts to increase the knowledge and tools for facilitating the discovery of such targets and pharmacologic agents directed against them should be prioritized.

Bleomycin reduces Vairimorpha (Nosema) ceranae infection in honey bees with some evident host toxicity.

Microsporidia cause disease in many beneficial insects, including honey bees, yet few pathogen control tools are available for protecting these important organisms against infection. Some evidence suggests that microsporidia possess a reduced number of genes encoding DNA repair proteins. We hypothesized that microsporidia would thus be susceptible to treatment with DNA-damaging agents and tested this hypothesis using a novel, rapid method for achieving robust and homogenous experimental infection of large numbers of newly emerged honey bees with one of its microsporidia pathogens, Vairimorpha (Nosema) ceranae. In carrying out these experiments, we found this novel V. ceranae inoculation method to have similar efficacy as other traditional methods. We show that the DNA-damaging agent bleomycin reduces V. ceranae levels, with minimal but measurable effects on honey bee survival and increased expression of midgut cellular stress genes, including those encoding SHSP. Increased expression of UpdlC suggests the occurrence of epithelial regeneration, which may contribute to host resistance to bleomycin treatment. While bleomycin does reduce infection levels, host toxicity issues may preclude its use in the field. However, with further work, bleomycin may provide a useful tool in the research setting as a potential selection agent for genetic modification of microsporidia.IMPORTANCEMicrosporidia cause disease in many beneficial insects, yet there are few tools available for control in the field or laboratory. Based on the reported paucity of DNA repair enzymes found in microsporidia genomes, we hypothesized that these obligate intracellular parasites would be sensitive to DNA damage. In support of this, we observed that the well-characterized DNA damage agent bleomycin can reduce levels of the microsporidia Vairimorpha (Nosema) ceranae in experimental infections in honey bees. Observation of slightly reduced honey bee survival and evidence of sublethal toxicity likely preclude the use of bleomycin in the field. However, this work identifies bleomycin as a compound that merits further exploration for use in research laboratories as a potential selection agent for generating genetically modified microsporidia.

Building-Level Detection Threshold of SARS-CoV-2 in Wastewater.

We established wastewater surveillance of SARS-CoV-2 in a small, residential, urban college as part of an integrated public health response during the COVID-19 pandemic. Students returned to campus in spring 2021. During the semester, students were required to perform nasal PCR tests twice weekly. At the same time, wastewater monitoring was established in 3 campus dormitory buildings. Two were dedicated dormitories with populations of 188 and 138 students; 1 was an isolation building where students were moved within 2 h of receiving positive test results. Analysis of wastewater from isolation indicated that the amount of viral shedding was highly variable and that viral concentration could not be used to estimate the number of cases at the building level. However, rapid movement of students to isolation enabled determination of predictive power, specificity, and sensitivity from instances in which generally one positive case at a time occurred in a building. Our assay yields effective results with an ~60% positive predictive power, ~90% negative predictive power, and ~90% specificity. Sensitivity, however, is low at ~40%. Detection is improved in the few instances of 2 simultaneous positive cases, with sensitivity of 1 case versus 2 cases increasing from ~20% to 100%. We also measured the appearance of a variant of concern on campus and noted a similarity in timeline with increased prevalence in surrounding New York City. Monitoring SARS-CoV-2 in the sewage outflow of individual buildings can be used with a realistic goal of containing outbreak clusters but not necessarily single cases. IMPORTANCE Diagnostic testing of sewage can detect levels of circulating viruses to help inform public health. Wastewater-based epidemiology has been particularly active during the COVID-19 pandemic to measure the prevalence of SARS-CoV-2. Understanding the technical limitations of diagnostic testing for individual buildings would help inform future surveillance programs. We report our diagnostic and clinical data monitoring of buildings on a college campus in New York City during the spring 2021 semester. Frequent nasal testing, mitigation measures, and public health protocols provided a context in which to study the effectiveness of wastewater-based epidemiology. Our efforts could not consistently detect individual positive COVID-19 cases, but sensitivity is significantly improved in detecting two simultaneous cases. We therefore contend that wastewater surveillance may be more practically suited for the mitigation of outbreak clusters.

Colonization of Honey Bee Digestive Tracts by Environmental Yeast Lachancea thermotolerans Is Naturally Occurring, Temperature Dependent, and Impacts the Microbiome of Newly Emerged Bees.

Honey bees are critical pollinators in both agricultural and ecological settings. Recent declines in honey bee colonies in the United States have put increased strain on agricultural pollination. Although there are many environmental stressors implicated in honey bee disease, there has been intensifying focus on the role of microbial attacks on honey bee health. Despite the long-standing appreciation for the association of fungi of various groups with honey bees and their broader environment, the effects of these interactions on honey bee health are incompletely understood. Here, we report the discovery of colonization of the honey bee digestive tract by the environmental yeast Lachancea thermotolerans. Experimental colonization of honey bee digestive tracts by L. thermotolerans revealed that this yeast species maintains high levels in the honey bee midgut only at temperatures below the typical colony temperature. In newly eclosed bees, L. thermotolerans colonization alters the microbiome, suggesting that environmental yeasts can impact its composition. Future studies should be undertaken to better understand the role of L. thermotolerans and other environmental yeasts in honey bee health. IMPORTANCE Although many fungal species are found in association with honey bees and their broader environment, the effects of these interactions on honey bee health are largely unknown. Here, we report the discovery that a yeast commonly found in the environment can be found at high levels in honey bee digestive tracts. Experimentally feeding this yeast to honey bees showed that the yeast's ability to maintain high levels in the digestive tract is influenced by temperature and can lead to alterations of the microbiome in young bees. These studies provide a foundation for future studies to better understand the role of environmental yeasts in honey bee health.

Paromomycin Reduces Vairimorpha (Nosema)ceranae Infection in Honey Bees but Perturbs Microbiome Levels and Midgut Cell Function.

Paromomycin is a naturally occurring aminoglycoside antibiotic that has effects on both prokaryotic and eukaryotic microbes. However, previous reports have indicated that it has little effect on microsporidia, including Vairimorpha (Nosema) ceranae, in cell culture models. V. ceranae is one of a number of microsporidia species that cause disease in honey bees and substantial efforts to find new treatment strategies for bees that are infected with these pathogens are ongoing. When testing compounds for potential activity against V. ceranae in whole organisms, we found that paromomycin reduces the infection intensity of this parasite. Critically, the necessary doses of paromomycin have high activity against the bacteria of the honey bee microbiome and cause evident stress in bees. Microsporidia have been shown to lack an essential binding site on the ribosome that is known to allow for maximal inhibition by paromomycin. Thus, it is possible that paromomycin impacts parasite levels through non-cell autonomous effects on microsporidia infection levels via effects on the microbiome or midgut cellular function. As paromomycin treatment could cause widespread honey bee health issues in agricultural settings, it does not represent an appropriate anti-microsporidia agent for use in the field.