Fecundity, in vitro early larval development and karyotype of the zoonotic nematode Anisakis pegreffii.

The in vitro life cycle of zoonotic helminths is an essential tool for -omic translational studies focused on disease control and treatment. Anisakiosis is an emerging zoonosis contracted by the ingestion of raw or undercooked fish infected with the third stage larvae (L3) of two sibling species Anisakis simplex sensu stricto (s.s.) and Anisakis pegreffii, the latter being the predominant species in the Mediterranean basin. Recently, in vitro culture of A. pegreffii has been developed to enable fast and large-scale production of fertile adults. However, the conditions for larval development from hatching to infective L3 were not fulfilled to complete the cycle. Herein, we used a Drosophila medium supplemented with chicken serum and adjusted different osmolarities to maintain the culture of L3 hatched from eggs for up to 17 weeks. The highest survival rate was observed in the medium with the highest osmolarities, which also allowed the highest larval exsheathment rate. Key morphological features of embryogenesis and postembryogenesis studied by transmission electron microscopy revealed that the excretory gland cell is differentiated already up to 48 h post-hatching. Extracellular vesicles and cell-free mitochondria are discharged between the two cuticle sheets of the second stage larvae (L2). Contemporarly cultivated, two populations of adult A. simplex s.s. and A. pegreffii reached an average production of 29,914.05 (± 27,629.36) and 24,370.96 (± 12,564.86) eggs/day/female, respectively. The chromosome spreads of A. pegreffii obtained from mature gonads suggests a diploid karyotype formula of 2n = 18. The development of a reliable protocol for the in vitro culture of a polyxenous nematode such as Anisakis spp. will serve to screen for much needed novel drug targets, but also to study the intricated and unknown ecological and physiological traits of these trophically transmitted marine nematodes.

Microsporidia in Commercially Harvested Marine Fish: A Potential Health Risk for Consumers.

Microsporidia are widely spread obligate intracellular fungal pathogens from vertebrate and invertebrate organisms, mainly transmitted by contaminated food and water. This study aims to detect the presence of major human-pathogenic microsporidia, i.e., Enterocytozoon bieneusi, Encephalitozoon intestinalis, Encephalitozoon hellem, and Encephalitozoon cuniculi, in the gastrointestinal tract of commercially harvested marine fish from Mediterranean coast of the Comunidad Valenciana, Eastern Spain. A total of 251 fish, 138 farmed fish and 113 wild fish from commercial fishing were tested by SYBR Green real-time PCR, enabling the simultaneous detection of the four targeted species. E. intestinalis/hellem was found in 1.45% of farmed fish and 7.96% of wild fish, while Enterocytozoonidae was detected in 2.90% and 18.58% of farmed and wild fish, respectively. E. cuniculi was not detected in any of the analyzed specimens. To the authors' knowledge, this is the first report of E. intestinalis/hellem in fish, particularly in marine fish. Although the role of fish in these species' epidemiology remains unknown, this finding points out a potential public health risk linked to fish consumption. Further studies are necessary to characterize these microsporidia in fish hosts better and to elucidate their epidemiological role.

Molecular Characterization of Cryptosporidium spp. in Cultivated and Wild Marine Fishes from Western Mediterranean with the First Detection of Zoonotic Cryptosporidium ubiquitum.

Fish not only harbor host-specific species/genotypes of Cryptosporidium, but also species like zoonotic C. parvum or anthroponotic C. hominis, which can pose a risk for fish consumers. This study aims to investigate fish cryptosporidiosis in an important aquaculture and fishery area of the Western Mediterranean (Comunidad Valenciana, Spain). We analyzed 404 specimens belonging to the following three groups: cultivated fish (N = 147), wild synanthropic fish (N = 147) and wild fish from extractive fisheries (N = 110). Nested PCR targeting the 18S rRNA gene, followed by sequencing and phylogenetic analysis, were performed. Positive isolates were also amplified at the actin gene locus. An overall prevalence of 4.2% was detected, with the highest prevalence in the synanthropic group (6.1%). C. molnari was identified in thirteen specimens from seven different host species. Zoonotic C. ubiquitum was detected in two European sea bass (Dicentrarchus labrax). One isolate similar to C. scophthalmi was detected in a cultivated meagre (Argyrosomus regius), and one isolate, highly divergent from all the Cryptosporidium species/genotypes described, was identified from a synanthropic round sardinella (Sardinella aurita). This study contributes to increasing the molecular data on fish cryptosporidiosis, expanding the range of known hosts for C. molnari and identifying, for the first time, zoonotic C. ubiquitum in edible marine fishes, pointing out a potential health risk.

Safe Game: Hygienic Habits in Self-Consumption of Game Meat in Eastern Spain.

We used anonymous questionnaires to assess the hygienic and sanitary aspects of game meat self-consumption in Eastern Spain as the first step towards a health risk assessment. The survey yielded 472 valid interviews from active hunters. The maximum possible score was 65 points (average 29 ± 8; range 1-52). Most participants were men (95%), but women achieved significantly better scores (p = 0.003). Hunters above 65 years old scored significantly lower results than younger groups (p = 0.007). The score increased with the educational level (p = 0.046). A 92% of the collaborators consumed game meat. Veterinary inspection and freezing were irregular among the participants. Most respondents declared carrying the animals in their personal vehicles. Of the dressing process, 61% of sites were outdoors, 68% of the participants declared using specific knives, 64% used the same clothes as in the field, and 42% used disposable gloves. The most usual way to dispose of the remains was garbage containers (41%); offal abandonment in the field was 33%, and 13% fed domestic animals using the remains. We conclude that public health authorities should increase their interest in the self-consumption of game meat. Clear guidelines about domestic dressing facilities and hygienic habits should be published, these being essential when looking for synergies with hunter associations.

Acute-to-Chronic Glycemic Ratio as a Predictor of COVID-19 Severity and Mortality.

OBJECTIVE: To evaluate the association between acute-to-chronic (A/C) glycemic ratio and mortality and severity outcomes for patients with type 2 diabetes (T2D) hospitalized with coronavirus disease 2019 (COVID-19). RESEARCH DESIGN AND METHODS: A total of 91 patients were included. We measured glycemia at admission and estimated the average chronic glucose levels to calculate the A/C glycemic ratio. The primary outcome was a composite of in-hospital mortality, intensive care unit admission, and mechanical ventilation. RESULTS: Thirty-five patients had a primary outcome event, presenting a significant association with the A/C glycemic ratio (hazard ratio [HR] 1.57 [95% CI 1.14-2.15], P = 0.005). In comparisons with the 2nd tertile, the 3rd tertile of the A/C glycemic ratio was associated with the primary outcome (HR 3.39 [95% CI 1.31-8.75], P = 0.012). In the multivariate analysis, after additional adjustment for age, sex, comorbidities, inflammatory markers, and corticosteroid therapy, the association for the 3rd tertile (HR 3.96 [95% CI 1.35-11.59], P = 0.012) remained significant. CONCLUSIONS: In patients with T2D hospitalized with COVID-19, the imbalance between acute glycemia at admission and chronic metabolic control is associated with worse prognosis.

Human and environmental factors driving Toxoplasma gondii prevalence in wild boar (Sus scrofa).

As one of the most relevant foodborne diseases, it is essential to know the factors related to the transmission, persistence and prevalence of Toxoplasma gondii infection. Eurasian wild boar (Sus scrofa) might play a relevant role in T.gondii's life cycle. This species is the most consumed big game animal in Spain and may act as a source of infection if the meat is eaten raw or undercooked or due to cross-contaminations. Additionally, wild boar can act as an excellent bioindicator of T.gondii circulation in the ecosystem, because its natural behaviour leads to exposure to oocysts from the soil when rooting and tissular bradyzoites when scavenging. A total of 1003 wild boar were sampled from 2010 to 2017 in Mediterranean Spain. Blood samples were tested with an indirect ELISA test giving a total of 14.1% (95% confidence interval 12.0-16.4%) positive results. The prevalence was not homogeneous in neither the animals nor the sampled districts. Significant differences were found regarding age, climatic conditions and human space occupancy. Human population aggregation, assessed by Demangeon's index, was identified as an influential factor in T.gondii infection risk. This multiple approach allows us to evaluate local risks for human and environmental contamination.

A comparative cost analysis between two quick diagnosis units of different levels of complexity.

Aim: To compare by micro-costing the costs incurred by quick diagnosis units of tertiary and second-level hospitals. Patients & methods: We included 407 patients from a tertiary and secondary hospital unit. A bottom-up approach was applied. Results: Cost per patient was €577.5 ± 219.6 in the tertiary versus €394.7 ± 92.58 in the secondary unit (p = 0.0559). Mean number of visits and ratio of successive/first visits were significantly higher in the former (3.098 and 2.07 vs 2.123 and 1.12, respectively). Personnel and indirect costs including their percent contribution to overall costs accounted for the main differences. Conclusion: A greater volume of appointments, number of staff and staff time and a greater complexity of patients from the tertiary hospital unit justified the differences in cost outcomes.

Potential Risk of Three Zoonotic Protozoa (Cryptosporidium spp., Giardia duodenalis, and Toxoplasma gondii) Transmission from Fish Consumption.

In recent decades, worldwide fish consumption has increased notably worldwide. Despite the health benefits of fish consumption, it also can suppose a risk because of fishborne diseases, including parasitic infections. Global changes are leading to the emergence of parasites in new locations and to the appearance of new sources of transmission. That is the case of the zoonotic protozoa Cryptosporidium spp., Giardia duodenalis, and Toxoplasma gondii; all of them reach aquatic environments and have been found in shellfish. Similarly, these protozoa can be present in other aquatic animals, such as fish. The present review gives an overview on these three zoonotic protozoa in order to understand their potential presence in fish and to comprehensively revise all the evidences of fish as a new potential source of Cryptosporidium spp., Giardia duodenalis, and Toxoplasma gondii transmission. All of them have been found in both marine and freshwater fishes. Until now, it has not been possible to demonstrate that fish are natural hosts for these protozoa; otherwise, they would merely act as mechanical transporters. Nevertheless, even if fish only accumulate and transport these protozoa, they could be a "new" source of infection for people.

Quick diagnosis units: predictors of time to diagnosis and costs.

Financial crisis has forced health systems to seek alternatives to hospitalization-based healthcare. Quick diagnosis units (QDUs) are cost-effective compared to hospitalization, but the determinants of QDU costs have not been studied.We aimed at assessing the predictors of costs of a district hospital QDU (Hospital Plató, Barcelona) between 2009 and 2016.This study was a retrospective longitudinal single center study of 404 consecutive outpatients referred to the QDU of Hospital Plató. The referral reason was dichotomized into suggestive of malignancy vs other. The final diagnosis was dichotomized into organic vs nonorganic and malignancy vs nonmalignancy. All individual resource costs were obtained from the finance department to conduct a micro-costing analysis of the study period.Mean age was 62 ±â€Š20 years (women = 56%), and median time-to-diagnosis, 12 days. Total and partial costs were greater in cases with final diagnosis of organic vs nonorganic disorder, as it was in those with symptoms suggestive or a final diagnosis of cancer vs noncancer. Of all subcosts, imaging showed the stronger correlation with total cost. Time-to-diagnosis and imaging costs were significant predictors of total cost above the median in binary logistic regression, with imaging costs also being a significant predictor in multiple linear regression (with total cost as quantitative outcome).Predictors of QDU costs are partly nonmodifiable (i.e., cancer suspicion, actually one of the goals of QDUs). Yet, improved primary-care-to-hospital referral circuits reducing time to diagnosis as well as optimized imaging protocols might further increase the QDU cost-effectiveness process. Prospective studies (ideally with direct comparison to conventional hospitalization costs) are needed to explore this possibility.

[Vitamin D deficiency and related factors in patients with osteoporotic hip fracture]. / Déficit de vitamina D en la fractura osteoporótica de cadera y factores asociados.

BACKGROUND AND OBJECTIVE: Hypovitaminosis D is frequent in the elderly, and it is especially prevalent among patients with hip fracture. The prevalence of vitamin D deficiency and its related factors are not well known in our population. The objective of this study was to determine the prevalence of hypovitaminosis D in patients with osteoporotic hip fracture and to analyze which factors are associated with this deficit. PATIENTS AND METHOD: Transversal study. Inclusion of all consecutive patients older than 65 years, admitted in our hospital with osteoporotic hip fracture during the period of March 2002-February 2003. The prevalence of hypovitaminosis D and secondary hyperparathyroidism were analysed. Sunlight exposure, functional and nutritional status, and presence of comorbidity were investigated. RESULTS: 324 patients were included. Mean (standard deviation) age was 83 (7) years, and 80% were female. Vitamin D deficiency was observed in 217 cases (67%; 95% confidence interval [CI], 62-72%); and 57% of these patients had secondary hyperparathyroidism. A low nutritional status -albumin < 4 g/l (odds ratio [OR] = 4.5; 95% CI, 1.3-16; p = 0.019)- and a low functional status (Barthel index < 60; OR = 3; 95% CI, 1.3-6.7; p = 0.008) - were factors independently associated with hypovitaminosis D. However, an active sunlight exposure was a protective factor (OR = 0.09; 95% CI, 0.02-0.5; p = 0.004). CONCLUSIONS: The prevalence of hypovitaminosis D is high in patients with osteoporotic hip fracture, and in more than a half of the cases a secondary hyperparathyroidism is observed. The vitamin D deficiency is especially prevalent among patients with low sunlight exposure and low nutritional and functional status.

Déficit de vitamina D en la fractura osteoporótica de cadera y factores asociados / Vitamin D deficiency and related factors in patients with osteoporotic hip fracture

Fundamento y objetivo: La hipovitaminosis D es frecuente entre los ancianos y especialmente prevalente en los pacientes con fractura de cadera. La prevalencia y los factores asociados a este déficit vitamínico son poco conocidos en nuestro medio. El objetivo del presente trabajo ha sido determinar la prevalencia de hipovitaminosis D en los pacientes con fractura osteoporótica de cadera y analizar los posibles factores que favorecen dicho déficit. Pacientes y método: Se ha realizado un estudio transversal, en el que se incluyó de forma consecutiva a todos los pacientes mayores de 65 años ingresados con fractura osteoporótica de cadera durante el período comprendido entre marzo de 2002 y febrero de 2003. Se investigó la prevalencia de hipovitaminosis D y de hiperparatiroidismo secundario. Se recogieron el grado de exposición solar, la capacidad funcional, el estado nutricional y la presencia de comorbilidad. Resultados: Se incluyó a 324 pacientes con una edad media (desviación estándar) de 83 (7) años; un 80% eran mujeres. Se observó hipovitaminosis D en 217 casos (67%; intervalo de confianza [IC] del 95%, 62-72%), de los que el 57% presentaba hiperparatiroidismo secundario. El mal estado nutricional ­valores de albúmina < 4 g/l (odds ratio [OR] = 4,5; IC del 95%, 1,3-16; p = 0,019)­ y la baja capacidad funcional ­índice de Barthel < 60 (OR = 3; IC del 95%, 1,3-6,7; p = 0,008)­ fueron factores que de forma independiente se asociaron a hipovitaminosis D, mientras que la exposición solar activa se mostró como un factor protector (OR = 0,09; IC del 95%, 0,02-0,5; p = 0,004). Conclusiones: La prevalencia de hipovitaminosis D es alta en los pacientes con una fractura osteoporótica de cadera y en más de la mitad de ellos condiciona la presencia de hiperparatiroidismo secundario. Este déficit es especialmente prevalente entre los individuos con poca exposición solar, mal estado nutricional y baja capacidad funcional

Background and objective: Hypovitaminosis D is frequent in the elderly, and it is especially prevalent among patients with hip fracture. The prevalence of vitamin D deficiency and its related factors are not well known in our population. The objective of this study was to determine the prevalence of hypovitaminosis D in patients with osteoporotic hip fracture and to analyze which factors are associated with this deficit. Patients and method: Transversal study. Inclusion of all consecutive patients older than 65 years, admitted in our hospital with osteoporotic hip fracture during the period of March 2002-February 2003. The prevalence of hypovitaminosis D and secondary hyperparathyroidism were analysed. Sunlight exposure, functional and nutritional status, and presence of comorbidity were investigated. Results: 324 patients were included. Mean (standard deviation) age was 83 (7) years, and 80% were female. Vitamin D deficiency was observed in 217 cases (67%; 95% confidence interval [CI], 62-72%); and 57% of these patients had secondary hyperparathyroidism. A low nutritional status ­albumin < 4 g/l (odds ratio [OR] = 4.5; 95% CI, 1.3-16; p = 0.019)­ and a low functional status (Barthel index < 60; OR = 3; 95% CI, 1.3-6.7; p = 0.008) ­ were factors independently associated with hypovitaminosis D. However, an active sunlight exposure was a protective factor (OR = 0.09; 95% CI, 0.02-0.5; p = 0.004). Conclusions: The prevalence of hypovitaminosis D is high in patients with osteoporotic hip fracture, and in more than a half of the cases a secondary hyperparathyroidism is observed. The vitamin D deficiency is especially prevalent among patients with low sunlight exposure and low nutritional and functional status