Caracterização molecular de Cryptosporidium em bezerros de assentamentos rurais da região Noroeste do estado de São Paulo, Brasil

The study was conducted on 25 properties of the settlements Sao Jose I and Salvador, located in the municipalities of Brejo Alegre and Birigui, in the State of Sao Paulo, Brazil. A record of variables was elaborated and included data such as gender, breed and age of the animals. A total of 231 stool samples were collected from bovines aged one to six months, 128 being females and 103 males, 131 crossbred and 100 Holstein. Among the 231 samples, 17 (7.36%) were positive for Crvptosporidium spp. both by malachite green negative staining and by nested-PCR. Of the 17 positive samples, 14 were sequenced in agarose gel. These sequences were detected between 99% and 100% of genetic similarity for the following species. One sequence was similar to C. parvum (AB513880.1), one to C. bovis (MF074602.1), two to C. ryanae (KT922233.1), one to C. felis (KM977642.1) and nine were similar for C. andersoni reference MF350628. C. andersoni was found in animals aged 2-6 months, an age group which is different from those described by several authors. The presence of C. parvum indicates that the calves in the studied region should be considered a potential source for zoonotic transmission. For the first time to our knowledge, C. felis was identified in cattle in America401491496FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP2012/13733-3O estudo foi realizado num total de 25 propriedades localizadas nos assentamentos São José I e Salvador, situados nos municípios de Brejo Alegre e Birigui, no estado de São Paulo. Um registro de variáveis foi elaborado, incluindo dados como sexo, raça e idade dos animais. Foram colhidas 231 amostras de fezes de bovinos de um a seis meses de idade, sendo 128 fêmeas e 103 machos, 131 mestiços e 100 da raça Holandesa. Entre os 231 bovinos examinados, 17 (7,36%) foram positivos para Cryptosporidium spp. tanto pela coloração negativa de verde malaquita como pela nested-PCR. Das 17 amostras positivas, 14 apresentaram amplificação pela eletroforese em gel de agarose suficiente para fazer o sequenciamento de DNA. Essas sequências foram detectadas similaridade genética entre 99% e 100% com as seguintes espécies. Uma sequência foi semelhante com C. parvum (referência: AB513880.1), uma com C. bovis(MF074602.1), duas com C. ryanae (KT922233.1), uma com C. felis (KM977642.1) e nove foram semelhantes com C. andersoni (MF350628). O estudo caracteriza a presença do Cryptosporidiumspp. em bovinos oriundos de propriedades produtoras de leite na região Noroeste do estado de São Paulo, sendo o C. andersoni a espécie mais prevalente nesses animais, principalmente em uma faixa etária diferente das descritas por diversos autores. A presença de C. parvum indica que os bezerros da região estudada devem ser considerados como uma fonte potencial de oocistos de espécies zoonóticas.Identificamos com ineditismo o C. felis em bovinos na América, o que corrobora outros estudos realizados na Polônia e Espanha e evidencia a presença de espécies de Cryptosporidium em fezes em hospedeiros não naturais. Palavras-chave: Bezerros. Caracterização molecular. Criptosporidiose. Nested-PCR.Protozoan of the genus Cryptosporidium are obligate intracellular coccidia with a cosmopolitan distribution. Biological data and various molecular genotyping tools indicate that the genus Cryptosporidium comprises about 30 valid species and more than 70 genotypes that can infect fishes, amphibians, reptiles, birds, and mammals (NAKAMURA; MEIRELES, 2015).Among the named species, 13 have been described in cattle. Of these, nine present zoonotic potential, being more prevalent in humans: Cryptosporidium hominis, Cryptosporidium parvum, Cryptosporidium meleagridis, Cryptosporidium cuniculus and, occasionally,Cryptosporidium muris, Cryptosporidium suis, Cryptosporidium ubiquitum, Cryptosporidium felis(BORNAY-LLINARES et al. 1999; CARDONA et al., 2015) and Cryptosporidium canis (KVÁČ et al., 2016).Cattle are mainly infected with C. parvum, Cryptosporidium bovis, Cryptosporidium ryanaeand Cryptosporidium andersoni (XIAO, 2010). Of these, only the first is of zoonotic importance, being a concern for public health, since human cryptosporidiosis can be acquired from infected calves (LEARMONTH et al., 2004; SOPWITH et al., 2005; DEL COCO et al., 2014). These four species of Cryptosporidium are frequently diagnosed in cattle of different ages. Among these, C. parvumis the most prevalent in young pre-weaning calves (less than two months old) and shows low host specificity, and some genotypes are considered to have high zoonotic potential (DIXON et al., 2011).During the post-weaning phase (three to 11 months of age) a reduction in the prevalence of C. parvum is observed, however, in this stage an increase of C. bovis and C. ryanae infections is noticed. The latter two species are not considered zoonotic and show high specificity for bovine. C. andersoni is the species that mainly infects post-weaned calves and adult cattle. These parasitic species can infect other ruminants and rarely humans (DIXON et al., 2011).Due to the ability of this protozoan to infect different hosts and its constant presence in the environment, humans can acquire the infection in several ways, such as through ingestion of food and water contaminated with oocysts, direct contact with infected people (anthroponotic) or animals (zoonotic). Increases in environmental contamination and infected reservoirs assist in zoonotic transmission, as human cryptosporidiosis is also caused by certain species that also affect ruminants. The contact of humans with the feces of these animals that contains oocysts or contaminated water constitutes routes of transmission to humans (DEL COCO et al., 2014

Follicular Histomorphometry and Evaluation of Ovarian Apoptosis in Queens of Different Age Groups

Background: In humans and bitchs the age is another factor that may affect the size of ovarian structures, verifying alterations in the quality of the pool and size of follicular structures, which can compromise the use of these structures for in vitro maturation. There are no reports correlating the morphometric characteristics of the follicles and ovarian apoptosis at different ages in cats. The aim of this study was to evaluate the histomorphometric parameters of follicular growth and the relationship with the occurrence of apoptosis in ovarian tissue of young, adult and senile queens.Materials, Methods & Results: Eighteen domestic queens, multiparous, of different breeds and age groups were used in this study and divided into three groups according to their ages: five months to one year - young; (7.8 ± 1.0 months); one to six years - adults (2.8 ± 0.5 years); and more than six years - senile (8.0 ± 0.9 years). Vaginal cytology was performed in order to characterize the estrous phase associated with plasma concentrations of progesterone. The morphology and percentage of the vaginal epithelium cells were evaluated and queens were classified into estrous and non-estrus and plasma concentrations of progesterone were determined. Ovarian samples were collected after ovariohysterectomy to routine histological processin and all follicles were counted and categorized into two groups, non-atresic and atresic. The mean follicular and oocyte diameters were calculated between the measurement of the largest diameter and perpendicular diameter. The relationship between follicle and oocyte were determined using the measurements of diameter, area and perimeter. The apoptotic cells were detected and cells were considered positive when TUNEL reaction was detected. The morphometric index of 1039 follicles were evaluated. Primordial follicles in young animals showed larger diameter, follicular area and perimeter than the structures of adult queens, as well as the unilaminar primary follicles of the same group were larger compared with senile animals (P 0.05). For the pre-ovulatory follicles there was no positive correlation between the oocyte growths regarding the follicles (P > 0.05). Only in senile animals positive markers for apoptosis were identified in nuclei of primordial follicles. No significant differences concerning the number of follicles and Tunel positive cells were observed between groups (P > 0.05).Discussion: Considering the importance of this study for greater knowledge in the basic aspects for reproductive biotechnologies, we verified that secondary follicles showed the largest diameters and younger animals the largest values for diameter, area and perimeter, suggesting that this age group could be ideal for the use and manipulation of oocytes. The process of follicular atresia is characterized by the occurrence of apoptosis, or programmed cell death when the organism begins to efficiently eliminate dysfunctional cells. The study of follicular apoptosis in small animals, especially in cats, is very important for the development of reproduction biotechnologies. Phenomenon of apoptosis showed no relationship with age in queens, occurring in a physiological, continuous and proportionate manner considering the number of nondominant follicles involved in each estrous cycle

The logic of bunched implications: a memoir

© 2006 Benjamin Robert Horsfall.This is a study of the semantics and proof theory of the logic of bunched implications (BI), which is promoted as a logic of (computational) resources, and is a foundational component of separation logic, an approach to program analysis. BI combines an additive, or intuitionistic, fragment with a multiplicative fragment. The additive fragment has full use of the structural rules of weakening and contraction, and the multiplicative fragment has none. Thus it contains two conjunctive and two implicative connectives. At various points, we illustrate a resource view of BI based upon the Kripke resource semantics. Our first original contribution is the formulation of a proof system for BI in the newly developed proof-theoretical formalism of the calculus of structures. The calculus of structures is distinguished by its employment of deep inference, but we already see deep inference in a limited form in the established proof theory for BI. We show that our system is sound with respect to the elementary Kripke resource semantics for BI, and complete with respect to a formulation of the partially-defined Kripke resource semantics. Our second contribution is the development from a semantic standpoint of preliminary ideas for a hybrid logic of bunched implications (HBI). We give a Kripke semantics for HBI in which nominal propositional atoms can be seen as names for resources, rather than as names for locations, as is the case with related proposals for BI-Loc and for intuitionistic hybrid logic

Follicular Histomorphometry and Evaluation of Ovarian Apoptosis in Queens of Different Age Groups

Background: In humans and bitchs the age is another factor that may affect the size of ovarian structures, verifying alterations in the quality of the pool and size of follicular structures, which can compromise the use of these structures for in vitro maturation. There are no reports correlating the morphometric characteristics of the follicles and ovarian apoptosis at different ages in cats. The aim of this study was to evaluate the histomorphometric parameters of follicular growth and the relationship with the occurrence of apoptosis in ovarian tissue of young, adult and senile queens.Materials, Methods & Results: Eighteen domestic queens, multiparous, of different breeds and age groups were used in this study and divided into three groups according to their ages: five months to one year - young; (7.8 ± 1.0 months); one to six years - adults (2.8 ± 0.5 years); and more than six years - senile (8.0 ± 0.9 years). Vaginal cytology was performed in order to characterize the estrous phase associated with plasma concentrations of progesterone. The morphology and percentage of the vaginal epithelium cells were evaluated and queens were classified into estrous and non-estrus and plasma concentrations of progesterone were determined. Ovarian samples were collected after ovariohysterectomy to routine histological processin and all follicles were counted and categorized into two groups, non-atresic and atresic. The mean follicular and oocyte diameters were calculated between the measurement of the largest diameter and perpendicular diameter. The relationship between follicle and oocyte were determined using the measurements of diameter, area and perimeter. The apoptotic cells were detected and cells were considered positive when TUNEL reaction was detected. The morphometric index of 1039 follicles were evaluated. Primordial follicles in young animals showed larger diameter, follicular area and perimeter than the structures of adult queens, as well as the unilaminar primary follicles of the same group were larger compared with senile animals (P 0.05). For the pre-ovulatory follicles there was no positive correlation between the oocyte growths regarding the follicles (P > 0.05). Only in senile animals positive markers for apoptosis were identified in nuclei of primordial follicles. No significant differences concerning the number of follicles and Tunel positive cells were observed between groups (P > 0.05).Discussion: Considering the importance of this study for greater knowledge in the basic aspects for reproductive biotechnologies, we verified that secondary follicles showed the largest diameters and younger animals the largest values for diameter, area and perimeter, suggesting that this age group could be ideal for the use and manipulation of oocytes. The process of follicular atresia is characterized by the occurrence of apoptosis, or programmed cell death when the organism begins to efficiently eliminate dysfunctional cells. The study of follicular apoptosis in small animals, especially in cats, is very important for the development of reproduction biotechnologies. Phenomenon of apoptosis showed no relationship with age in queens, occurring in a physiological, continuous and proportionate manner considering the number of nondominant follicles involved in each estrous cycle

Relevant aspects of human toxoplasmosis

Considering the great importance that the infection by Toxoplasma gondii has in public health, the aim of this article was to demonstrate some aspects about this disease in human being. This parasite can cause several behavioral changes and many cases of reactivation of this disease are associated with Acquired Immunodeficiency Syndrome – AIDS. Major cases of human infections by T. gondii are assymptomatic and the main clinical manifestation in the acute phase of this zoonosis is lymphadenopathy, usually in the cervical area. There may also be fever, night sweats, myalgia, maculopapular rash lymphocytic atypia and chorioretinitis. The congenital and/or acquired chronic latent cerebral toxoplasmosis play an important role in the development of some types of neurodegeneration, like Alzheimer disease and Down Syndrome. Humans and animals should not consume raw or undercooked meat, unpasteurized or not boiled dairy products, water without being treated and poorly washed fruits and vegetables. Educational programs focused on reducing T. gondii environmental contamination are essential for the congenital infection control. Due to the severity of this kind of infection and its sequel, it is critical that those responsible for the community’s health establish prevention programs to avoid such a situation, obtaining results in the medium and long term, and particularly preserving the involved population’s welfare. A lack or incomplete prenatal treatment was identified as an important risk factor for congenital toxoplasmosis, reinforcing the need of improvement of prenatal care. In this study was demonstrated the importance of implementing prevention programs to guide pregnant women to prevent the infection. Preventive measures like information and health education; screening of pregnant women and infants; limiting harm from risk behaviour; treatment of risk cases and vaccination are recommended. Serum monitoring throughout pregnancy, so as detect cases of maternal seroconversion allowing for early maternal treatment is also basic. Some countries have adopted control programs of human toxoplasmosis, however, this issue should be of world attention, considering its relevant aspects of public health, requiring the establishment of community education campaigns and monitoring of pregnant women in the pre and neonatal, especially in a preventive action to reduce the pathogenic effects of this disease.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP