Isolation of Ovicidal Fungi from Fecal Samples of Captive Animals Maintained in a Zoological Park

There are certain saprophytic fungi in the soil able to develop an antagonistic effect against eggs of parasites. Some of these fungal species are ingested by animals during grazing, and survive in their feces after passing through the digestive tract. To identify and isolate ovicidal fungi in the feces of wild captive animals, a total of 60 fecal samples were taken from different wild animals kept captive in the Marcelle Natureza Zoological Park (Lugo, Spain). After the serial culture of the feces onto Petri dishes with different media, their parasicitide activity was assayed against eggs of trematodes (Calicophoron daubneyi) and ascarids (Parascaris equorum). Seven fungal genera were identified in the feces. Isolates from Fusarium, Lecanicillium, Mucor, Trichoderma, and Verticillium showed an ovicidal effect classified as type 3, because of their ability to adhere to the eggshell, penetrate, and damage permanently the inner embryo. Penicillium and Gliocladium developed a type 1 effect (hyphae attach to the eggshell but morphological damage was not provoked). These results provide very interesting and useful information about fungi susceptible for being used in biological control procedures against parasitesThis research was partly supported by the Research Projects AGL2012-34355 and CTM2015-65954-R (Ministerio de Economía y Competitividad, Spain; FEDER). María S. Arias is recipient of a “Parga Pondal” postdoctoral research fellowship (Xunta de Galicia, Spain)S

The Control of Zoonotic Soil-Transmitted Helminthoses Using Saprophytic Fungi

Soil-transmitted helminths (STHs) are parasites transmitted through contact with soil contaminated with their infective eggs/larvae. People are infected by exposure to human-specific species or animal species (zoonotic agents). Fecal samples containing eggs of Ascaris suum or Lemurostrongylus sp. were sprayed with spores of the soil saprophytic filamentous fungi Clonostachys rosea (CR) and Trichoderma atrobrunneum (TA). The antagonistic effect was assessed by estimating the viability of eggs and their developmental rate. Compared to the controls (unexposed to fungi), the viability of the eggs of A. suum was halved in CR and decreased by two thirds in TA, while the viability of the eggs of Lemurostrongylus sp. was reduced by one quarter and one third in CR and TA treatments, respectively. The Soil Contamination Index (SCI), defined as the viable eggs that attained the infective stage, reached the highest percentages for A. suum in the controls after four weeks (66%), with 21% in CL and 11% in TA. For Lemurostrongylus sp., the values were 80%, 49%, and 41% for control, CR and TA treatments, respectively. We concluded that spreading spores of C. rosea or T. atrobrunneum directly onto the feces of animal species represents a sustainable approach under a One Health context to potentially reduce the risk of zoonotic STHs in humansThis research was funded by the Research Projects CTM2015-65954-R and RYC-2016-21407 (Ministerio de Economía y Competitividad, Spain; FEDER), and ED431F 2018/03 (Consellería de Cultura, Educación e Ordenación Universitaria, Xunta de Galicia, Spain). María Sol Arias Vázquez is recipient of a Ramón y Cajal (Ministerio de Economía y Competitividad, Spain) contractS

Mixed Production of Filamentous Fungal Spores for Preventing Soil-Transmitted Helminth Zoonoses: A Preliminary Analysis

Helminth zoonoses are parasitic infections shared by humans and animals, being the soil-transmitted helminths (STHs) mainly caused by roundworms (ascarids) and hookworms. This study was aimed to assess the individual and/or mixed production of two helminth-antagonistic fungi, one ovicide (Mucor circinelloides) and other predator (Duddingtonia flagrans). Fungi were grown both in Petri plates and in a submerged culture (composed by water, NaCl, Na2HPO4 · 12 H2O, and wheat (Triticum aestivum)). A Fasciola hepatica recombinant protein (FhrAPS) was incorporated to the cultures to improve fungal production. All the cultured plates showed fungal growth, without difference in the development of the fungi when grown alone or mixed. High counts of Mucor spores were produced in liquid media cultures, and no significant differences were achieved regarding single or mixed cultures, or the incorporation of the FhrAPS. A significantly higher production of Duddingtonia spores after the incorporation of the FhrAPS was observed. When analyzing the parasiticide efficacy of the fungal mixture, viability of T. canis eggs reduced to 51%, and the numbers of third stage cyathostomin larvae reduced to 4%. It is concluded, the capability of a fungal mixture containing an ovicide (Mucor) and a predator species (Duddingtonia) for growing together in a submerged medium containing the FhrAPS offers a very interesting tool for preventing STHs.This work was partly supported by the Research Projects 10MDS261023PR (Xunta de Galicia, Spain) and AGL2012-34355 (Ministerio de Economía y Competitividad, Spain; FEDER)S

Advantageous Fungi against Parasites Transmitted through Soil

Although many fungal specimens are responsible for human and/or animal infection, other species are advantageous for preventing the infection by soil-transmitted zoonotic parasites. Infection occurs by the accidental ingestion of parasitic stages (cysts, oocysts, eggs, and larvae), their active penetration through the skin or through direct contact. Numerous species of helminths develop an external phase in the soil where the infective stages are attained, thus mammals become infected when grazing, drinking, or accidentally. Ectoparasites as ticks perform also in the soil the phase from egg to larva. Different soil saprophytic fungi that turn into predatory agents when parasitic stages are near have been isolated and described. These species are capable of destroying the pathogens or irreversibly decreasing their viability, providing thus a very interesting and sustainable tool to reduce environmental contamination by pathogenic agents. In the last year, a profound knowledge on the most appropriate fungal species, together with the proper way to disseminate them, has been acquired

Prevalence and distribution of infectious and parasitic agents in roe deer from Spain and their possible role as reservoirs

In order to provide up-to-date information about the prevalence of infectious and parasitic agents in Spanish roe deer (Capreolus capreolus), samples from 93 animals hunted from January 2013 to April 2015 were collected and analysed by parasitological, serological and molecular techniques. Sampled animals came from four roe deer populations corresponding to Oceanic, Continental, Mediterranean and Mountainous ecosystems of Spain. Data regarding sex, age and year were also considered. A high percentage of roe deer (95.7%) resulted positive for at least one agent. Sarcocystis spp. was the most frequently diagnosed genus (88.8%), followed by gastrointestinal nematodes (62.9%) and Schmallenberg virus (53.5%). Varestrongylus capreoli (38%), Anaplasma phagocitophylum (34.2%), Eimeria spp. (29.2%), Toxoplasma gondii (25%) and Cephenemyia stimulator (23.8%) displayed medium prevalences and, finally low percentages were registered for Moniezia spp. (6.7%), Dictyocaulus noerneri (2.4%) and Mycobacterium avium subsp. paratuberculosis (1.5%). No infections by Neospora caninum, Bovine herpesvirus, pestivirus or Coxiella burnetii were found. Climate was significantly associated with the prevalence of T. gondii, C. stimulator and A. phagocitophylum, with higher prevalences in animals from Oceanic and Mediterranean areas. Our results suggest that infections affecting Spanish roe deer, especially those of parasitic aetiology, represent one of the causes of the descent in the abundance of this ungulate in the last years. Moreover, the high prevalence of zoonotic agents such as T. gondii and A. phagocitophylum could also have a great relevance in the environmental and/or Public Health.This research was supported by a Programme for consolidating and structuring competitive research groups (GRC2015/003, Xunta de Galicia) and by the Research Project ‘RUMIGAL: Rede de estudo multidisciplinar dos ruminantes en Galicia’ (R2014/005, Xunta de Galicia, Spain)S

Implementation of Biological Control to the Integrated Control of Strongyle Infection among Wild Captive Equids in a Zoological Park

The integrated control of strongyles was assayed for a period of three years in wild equids (zebras, European donkeys, and African wild asses) captive in a zoo and infected by strongyles. During three years control of parasites consisted of deworming with ivermectin + praziquantel; equids also received every two days commercial nutritional pellets containing a blend of 104 - 105 spores of the fungi Mucor circinelloides + Duddingtonia flagrans per kg meal. Coprological analyses were done monthly to establish the counts of eggs of strongyles per gram of feces (EPG). The reductions in the fecal egg counts (FECR) and in the positive horses (PHR) were calculated fifteen days after deworming; the egg reappearance period (ERP) and the time elapsed from the previous deworming (TPD) were also recorded. Four anthelmintic treatments were administered during the assay, three times throughout the first 2 yrs, and another treatment during the last one. FECR values of 96-100% and 75-100% for the PHR were recorded. The ERP oscillated between eight and twenty-eight weeks, and the TPD ranged from four to eighteen months, increasing to the end of the trial. No side effects were observed in any of the equids. It is concluded that integrated control of strongyles among equids captive in a zoo can be developed by anthelmintic deworming together with the administration of pellets manufactured with spores of parasiticide fungi every two days

A Novel second instar <i>Gasterophilus</i> excretory/secretory antigen-based ELISA for the diagnosis of gasterophilosis in grazing horses

We have developed a novel enzyme-linked immunosorbent assay (ELISA) based on excretory/secretory antigens of second instar Gasterophilus for the diagnosis of gasterophilosis in grazing horses. Between January 2007 and January 2009, two experiments were carried out on free-ranging horses in northwest Spain. During the first year, monthly blood samples were collected from a herd of 25 horses. In the second year, a monthly serological survey was conducted for a total of 398 different horses. All the sera were analyzed by ELISA using excretory/secretory antigens from Gasterophilus intestinalis (GphiL2ES) and Gasterophilus nasalis second-stage larvae (GphnL2ES). Climatic data were collected between January 2007 and January 2009 from local meteorological automated stations to establish the weather pattern in the study area. Observations of Gasterophilus eggs on the horses’ hair and third instars passed in the faeces were also done. The kinetics of IgG response decreased against GphiL2ES from January to July, increased slowly from August and rose up to January. After a slight decrease in January, the absorbances against GphnL2ES reduced from April to August, when the lowest values were observed. The IgG values rose until the end of the study in January. Third instars were observed in the faeces in March to May, and Gasterophilus eggs were seen on the horses’ hair from June to September. The highest IgG seroprevalences were achieved in winter (January–February; 100%) against both antigens. The lowest percentages of seropositivity were observed in June (3%) to the GphiL2ES, and in July (9%) to the GphnL2ES. The use of antigens from G. intestinalis second-stage larvae was shown to be suitable for diagnosing infestation by G. intestinalis or G. nasalis. We concluded that under oceanic climate conditions, the egg-laying period occurs from late spring, and eggs and first instars are found in the mouth in early summer. During summer the second instars move into the stomach and intestine, where the third-stage larvae remain until the end of winter, when pupation takes place. The adult horse bot fly emerges in the spring. Two treatments for the control of gasterophilosis are suggested: a curative in the summer to eliminate the first instars and a preventive in the autumn to suppress the second instars

Effects of pre-operative isolation on postoperative pulmonary complications after elective surgery: an international prospective cohort study an international prospective cohort study

We aimed to determine the impact of pre-operative isolation on postoperative pulmonary complications after elective surgery during the global SARS-CoV-2 pandemic. We performed an international prospective cohort study including patients undergoing elective surgery in October 2020. Isolation was defined as the period before surgery during which patients did not leave their house or receive visitors from outside their household. The primary outcome was postoperative pulmonary complications, adjusted in multivariable models for measured confounders. Pre-defined sub-group analyses were performed for the primary outcome. A total of 96,454 patients from 114 countries were included and overall, 26,948 (27.9%) patients isolated before surgery. Postoperative pulmonary complications were recorded in 1947 (2.0%) patients of which 227 (11.7%) were associated with SARS-CoV-2 infection. Patients who isolated pre-operatively were older, had more respiratory comorbidities and were more commonly from areas of high SARS-CoV-2 incidence and high-income countries. Although the overall rates of postoperative pulmonary complications were similar in those that isolated and those that did not (2.1% vs 2.0%, respectively), isolation was associated with higher rates of postoperative pulmonary complications after adjustment (adjusted OR 1.20, 95%CI 1.05–1.36, p = 0.005). Sensitivity analyses revealed no further differences when patients were categorised by: pre-operative testing; use of COVID-19-free pathways; or community SARS-CoV-2 prevalence. The rate of postoperative pulmonary complications increased with periods of isolation longer than 3 days, with an OR (95%CI) at 4–7 days or ≥ 8 days of 1.25 (1.04–1.48), p = 0.015 and 1.31 (1.11–1.55), p = 0.001, respectively. Isolation before elective surgery might be associated with a small but clinically important increased risk of postoperative pulmonary complications. Longer periods of isolation showed no reduction in the risk of postoperative pulmonary complications. These findings have significant implications for global provision of elective surgical care. We aimed to determine the impact of pre-operative isolation on postoperative pulmonary complications after elective surgery during the global SARS-CoV-2 pandemic. We performed an international prospective cohort study including patients undergoing elective surgery in October 2020. Isolation was defined as the period before surgery during which patients did not leave their house or receive visitors from outside their household. The primary outcome was postoperative pulmonary complications, adjusted in multivariable models for measured confounders. Pre-defined sub-group analyses were performed for the primary outcome. A total of 96,454 patients from 114 countries were included and overall, 26,948 (27.9%) patients isolated before surgery. Postoperative pulmonary complications were recorded in 1947 (2.0%) patients of which 227 (11.7%) were associated with SARS-CoV-2 infection. Patients who isolated pre-operatively were older, had more respiratory comorbidities and were more commonly from areas of high SARS-CoV-2 incidence and high-income countries. Although the overall rates of postoperative pulmonary complications were similar in those that isolated and those that did not (2.1% vs 2.0%, respectively), isolation was associated with higher rates of postoperative pulmonary complications after adjustment (adjusted OR 1.20, 95%CI 1.05–1.36, p = 0.005). Sensitivity analyses revealed no further differences when patients were categorised by: pre-operative testing; use of COVID-19-free pathways; or community SARS-CoV-2 prevalence. The rate of postoperative pulmonary complications increased with periods of isolation longer than 3 days, with an OR (95%CI) at 4–7 days or ≥ 8 days of 1.25 (1.04–1.48), p = 0.015 and 1.31 (1.11–1.55), p = 0.001, respectively. Isolation before elective surgery might be associated with a small but clinically important increased risk of postoperative pulmonary complications. Longer periods of isolation showed no reduction in the risk of postoperative pulmonary complications. These findings have significant implications for global provision of elective surgical care