Microbiome Structure of a Wild Drosophila Community along Tropical Elevational Gradients and Comparison to Laboratory Lines.

Variation along environmental gradients in host-associated microbial communities is not well understood compared to free-living microbial communities. Because elevational gradients may serve as natural proxies for climate change, understanding patterns along these gradients can inform our understanding of the threats hosts and their symbiotic microbes face in a warming world. In this study, we analyzed bacterial microbiomes from pupae and adults of four Drosophila species native to Australian tropical rainforests. We sampled wild individuals at high and low elevations along two mountain gradients to determine natural diversity patterns. Further, we sampled laboratory-reared individuals from isofemale lines established from the same localities to see if any natural patterns are retained in the lab. In both environments, we controlled for diet to help elucidate other deterministic patterns of microbiome composition. We found small but significant differences in Drosophila bacterial community composition across elevation, with some notable taxonomic differences between different Drosophila species and sites. Further, we found that field-collected fly pupae had significantly richer microbiomes than laboratory-reared pupae. We also found similar microbiome composition in both types of provided diet, suggesting that the significant differences found among Drosophila microbiomes are the products of surrounding environments with different bacterial species pools, possibly bound to elevational differences in temperature. Our results suggest that comparative studies between lab and field specimens help reveal the true variability in microbiome communities that can exist within a single species. IMPORTANCE Bacteria form microbial communities inside most higher-level organisms, but we know little about how the microbiome varies along environmental gradients and between natural host populations and laboratory colonies. To explore such effects on insect-associated microbiomes, we studied the gut microbiome in four Drosophila species over two mountain gradients in tropical Australia. We also compared these data to individuals kept in the laboratory to understand how different settings changed microbiome communities. We found that field-sampled individuals had significantly higher microbiome diversity than those from the lab. In wild Drosophila populations, elevation explains a small but significant amount of the variation in their microbial communities. Our study highlights the importance of environmental bacterial sources for Drosophila microbiome composition across elevational gradients and shows how comparative studies help reveal the true flexibility in microbiome communities that can exist within a species.

DROP: Molecular voucher database for identification of Drosophila parasitoids.

Molecular identification is increasingly used to speed up biodiversity surveys and laboratory experiments. However, many groups of organisms cannot be reliably identified using standard databases such as GenBank or BOLD due to lack of sequenced voucher specimens identified by experts. Sometimes a large number of sequences are available, but with too many errors to allow identification. Here, we address this problem for parasitoids of Drosophila by introducing a curated open-access molecular reference database, DROP (Drosophila parasitoids). Identifying Drosophila parasitoids is challenging and poses a major impediment to realize the full potential of this model system in studies ranging from molecular mechanisms to food webs, and in biological control of Drosophila suzukii. In DROP, genetic data are linked to voucher specimens and, where possible, the voucher specimens are identified by taxonomists and vetted through direct comparison with primary type material. To initiate DROP, we curated 154 laboratory strains, 856 vouchers, 554 DNA sequences, 16 genomes, 14 transcriptomes, and six proteomes drawn from a total of 183 operational taxonomic units (OTUs): 114 described Drosophila parasitoid species and 69 provisional species. We found species richness of Drosophila parasitoids to be heavily underestimated and provide an updated taxonomic catalogue for the community. DROP offers accurate molecular identification and improves cross-referencing between individual studies that we hope will catalyse research on this diverse and fascinating model system. Our effort should also serve as an example for researchers facing similar molecular identification problems in other groups of organisms.

Effects of selected Indonesian plant extracts on E. cuniculi infection in vivo.

The present study was conducted to evaluate the methanolic extracts from several plant leaves widely used in traditional medicine to cure digestive tract disorders and in the self-medication of wild animals such as non-human primates, namely Archidendron fagifolium, Diospyros sumatrana, Shorea sumatrana, and Piper betle leaves, with regard to their antimicrosporidial activity against Encephalitozoon cuniculi in immunocompetent BALB/c mice determined using molecular detection of microsporidial DNA (qPCR) in various tissues and body fluids of infected, treated mice. Of the plant extracts tested, Diospyros sumatrana provided the most promising results, reducing spore shedding by 88% compared to untreated controls. Moreover, total burden per 1 g of tissue in the D. sumatrana extract-treated group reached 87% reduction compared to untreated controls, which was comparable to the effect of the standard drug, Albendazole. This data represents the baseline necessary for further research focused on determining the structure, activity and modes of action of the active compounds, mainly of D. sumatrana, enabling subsequent development of antimicrosporidial remedies.

First description of Cryptosporidium ubiquitum XIIa subtype family in farmed fur animals.

This study investigated the prevalence of Cryptosporidium in farmed fur animals in the Czech Republic and Poland. A total of 480 faecal samples were collected from fur animals, including 300 American mink (Mustela vison), 60 silver foxes (Vulpes vulpes), 50 long-tailed chinchillas (Chinchilla lanigera), and 70 nutrias (Myocastor coypus), at 14 farms. Samples were examined for the presence of Cryptosporidium using microscopy (following aniline-carbol-methyl violet staining) and sequence analysis of PCR amplified products. Three mink and two chinchillas from two different farms tested positive for Cryptosporidium ubiquitum DNA. The presence of C. ubiquitum DNA was not associated with diarrhoea. Subtyping of C. ubiquitum isolates by sequence analysis of the 60-kDa glycoprotein gene showed that isolates belonged to the XIIa subtype family, which was previously restricted to humans and ruminants. This suggests that C. ubiquitum subtype XIIa has a broader host range than previously reported.

Energy parasites trigger oncogene mutation.

PURPOSE: Cancer initialization can be explained as a result of parasitic virus energy consumption leading to randomized genome chemical bonding. MATERIALS AND METHODS: Analysis of experimental data on cell-mediated immunity (CMI) containing about 12,000 cases of healthy humans, cancer patients and patients with precancerous cervical lesions disclosed that the specific cancer and the non-specific lactate dehydrogenase-elevating (LDH) virus antigen elicit similar responses. The specific antigen is effective only in cancer type of its origin but the non-specific antigen in all examined cancers. CMI results of CIN patients display both healthy and cancer state. The ribonucleic acid (RNA) of the LDH virus parasitizing on energy reduces the ratio of coherent/random oscillations. Decreased effect of coherent cellular electromagnetic field on bonding electrons in biological macromolecules leads to elevating probability of random genome reactions. RESULTS: Overlapping of wave functions in biological macromolecules depends on energy of the cellular electromagnetic field which supplies energy to bonding electrons for selective chemical bonds. CMI responses of cancer and LDH virus antigens in all examined healthy, precancerous and cancer cases point to energy mechanism in cancer initiation. CONCLUSIONS: Dependence of the rate of biochemical reactions on biological electromagnetic field explains yet unknown mechanism of genome mutation.

Diseases caused by defects of energy level and loss of coherence in living cells.

Human and animal diseases are brought about by pathological alterations of production, composition, and conformation of macromolecules and structures in cells. Additional contributing factors include changes in physiological states caused by disturbances of energy supply, energy transduction, energy dissipation in moving or oscillating parts, and parasitic energy consumption. Disturbances of energy states may endanger existence of the system. The cell-mediated immunity (CMI) response of T lymphocytes correlating with their adherence properties was examined using antigen prepared from the serum of inbred laboratory mice strain C3H H(2k) infected with lactate dehydrogenase elevating (LDH) virus. LDH virus is a parasite on the cellular energy system. Significant CMI response was elicited in T lymphocytes prepared from the blood of patients with cancer of different phenotypes, acute myocardial infarctions, schizophrenia, and recurrent spontaneous abortions in early pregnancy from unknown reasons. The CMI response is assumed to monitor transferred information about decreased levels of energy states and decoherence in the cells caused by mitochondrial malfunction, parasitic consumption, production of lactate, and possibly other disturbances. The LDH virus infection or similar pathological processes caused by different agents might be connected with the diseases and monitored by the examined CMI response. A large amount of mitoses with chromosome defects in aborted fetuses suggest increased mutability of genomes caused by defective energy states.

Targeting mitochondria for cancer treatment - two types of mitochondrial dysfunction.

Two basic types of cancers were identified – those with the mitochondrial dysfunction in cancer cells (the Warburg effect) or in fibroblasts supplying energy rich metabolites to a cancer cell with functional mitochondria (the reverse Warburg effect). Inner membrane potential of the functional and dysfunctional mitochondria measured by fluorescent dyes (e.g. by Rhodamine 123) displays low and high values (apparent potential), respectively, which is in contrast to the level of oxidative metabolism. Mitochondrial dysfunction (full function) results in reduced (high) oxidative metabolism, low (high) real membrane potential, a simple layer (two layers) of transported protons around mitochondria, and high (low) damping of microtubule electric polar vibrations. Crucial modifications are caused by ordered water layer (exclusion zone). For the high oxidative metabolism one proton layer is at the mitochondrial membrane and the other at the outer rim of the ordered water layer. High and low damping of electric polar vibrations results in decreased and increased electromagnetic activity in cancer cells with the normal and the reverse Warburg effect, respectively. Due to nonlinear properties the electromagnetic frequency spectra of cancer cells and transformed fibroblasts are shifted in directions corresponding to their power deviations resulting in disturbances of interactions and escape from tissue control. The cancer cells and fibroblasts of the reverse Warburg effect tumors display frequency shifts in mutually opposite directions resulting in early generalization. High oxidative metabolism conditions high aggressiveness. Mitochondrial dysfunction, a gate to malignancy along the cancer transformation pathway, forms a narrow neck which could be convenient for cancer treatment.

Is the small size of a breast cancer tumor the crucial point for successful medical treatment?

The presented case displays a clinical study of a cancer phenotype with a poor clinical outcome. Prediction of cancer development and effects of treatment at the beginning of the clinical stage is difficult as the knowledge of cancer process and all necessary parameters of the host body are limited. Cancer is mainly studied on the basis of biochemical-genetic processes and their morphological manifestation. However, the malignant process is assumed to be of essential biophysical nature and develops after mitochondrial dysfunction, which is a direct result of oncogene mutation. Cancers based on the normal and the reverse Warburg effect should be distinguished. The cancer tumors with the reverse Warburg effect display aggressiveness associated with a high rate of recurrence and metastatic implantation. Besides the nature of the two basic types of breast cancer tumors the outcome depends not only on their type, size, and site but also on reactions and interaction with the surrounding tissue and the body aptitude for metastatic activity connected with individual blood or lymphatic vessels for metastatic transport. It is necessary to assess all favourable and adverse factors for cancer development. General reliable method of their specification for all cancers is not available. Nevertheless, the main factor seems to be aggressiveness of cancer cells as follows from interpretation. To reveal the aggressive reverse Warburg effect tumors, metabolic biomarkers of the fibroblast stress should be examined.

Biophysical insights into cancer transformation and treatment.

Biological systems are hierarchically self-organized complex structures characterized by nonlinear interactions. Biochemical energy is transformed into work of physical forces required for various biological functions. We postulate that energy transduction depends on endogenous electrodynamic fields generated by microtubules. Microtubules and mitochondria colocalize in cells with microtubules providing tracks for mitochondrial movement. Besides energy transformation, mitochondria form a spatially distributed proton charge layer and a resultant strong static electric field, which causes water ordering in the surrounding cytosol. These effects create conditions for generation of coherent electrodynamic field. The metabolic energy transduction pathways are strongly affected in cancers. Mitochondrial dysfunction in cancer cells (Warburg effect) or in fibroblasts associated with cancer cells (reverse Warburg effect) results in decreased or increased power of the generated electromagnetic field, respectively, and shifted and rebuilt frequency spectra. Disturbed electrodynamic interaction forces between cancer and healthy cells may favor local invasion and metastasis. A therapeutic strategy of targeting dysfunctional mitochondria for restoration of their physiological functions makes it possible to switch on the natural apoptotic pathway blocked in cancer transformed cells. Experience with dichloroacetate in cancer treatment and reestablishment of the healthy state may help in the development of novel effective drugs aimed at the mitochondrial function.

The impact of lower induction values of 50 Hz external electromagnetic fields on in vitro T lymphocyte adherence capabilities.

Our research thus far has concerned the impact of external electromagnetic fields (50 Hz) and low (0.01-10 mT) induction on adherence capabilities of T lymphocytes obtained from the blood of patients with head and neck tumors. We know that the in vitro adherence capability of T lymphocytes towards surfaces in cancer patients is less than that of control. Previously, we have found that exposure to electromagnetic fields (50 Hz/0.01-10 mT) increases the capability of T lymphocytes, in larynx/pharynx cancer patients, to adhere in vitro to surfaces, achieving almost physiological values, in not only pre-treatment patients but also those receiving treatment in the course of follow-up. The capability of T lymphocytes in controls (voluntary blood donors) to adhere to surfaces was also increased (50 Hz/0.01-0.5 mT). The present study concentrates on the significance of the level of electromagnetic field induction in order to determine whether low induction values can restore T lymphocytes adherence capabilities. Testing a subset of 20 patients showed a statistically significant difference (p<0.05) in the in vitro adherence capacity of T lymphocytes between both 0.01 and 0.05, and 0.1 mT induction levels. In the control group (patients diagnosed with chronic sensorineural hearing loss) there was even a statistically significant difference between induction values of 0.05 and 0.01 mT. A statistically significant difference (p<0.05) was also achieved with induction levels of 1 and 10 mT compared to 0.5, 0.1, and 0.05 mT, respectively. Therefore, we concluded that lower induction values resulted in a more biologically significant response.