Norovirus and human adenovirus occurrence and diversity in recreational water in a karst aquifer in the Yucatan Peninsula, Mexico.

AIMS: To determine the seasonal occurrence and diversity of norovirus (NoV) and human adenovirus (HAdV) in groundwater from sinkholes, and brackish water used for recreational activities in the karst aquifer of the Yucatan Peninsula, Mexico. METHODS AND RESULTS: Hollow fibre ultrafiltration was used to concentrate viruses and standard plaque assay methods were used to enumerate somatic and F+ specific coliphages as viral indicators. Real-time quantitative polymerase chain reaction assays were used to estimate the number of genome copies for NoV strains GI, and GII, and HAdVs. The predominant NoV genotypes and HAdV serotypes were identified by comparative sequence analysis. Somatic and male F+ specific coliphages were detected at concentrations up to 94 and 60 plaque-forming units per 100 ml respectively. The NoV genogroup I (GI) was associated with 50% of the sampled sites during the rainy season only, at concentrations ranging from 120 to 1600 genome copies per litre (GC l-1 ). The NoV genogroup II (GII) was detected in 30 and 40% of the sampled sites during the rainy and dry seasons, respectively, at concentrations ranging from 10 to 290 GC l-1 . During the rainy and dry seasons, HAdVs were detected in 20% of the sites, at concentrations ranging from 24 to 690 GC l-1 . Identification of viral types revealed the presence of NoV GI.2, GII.Pe, GII.P16 and GII.P17, and HAdV F serotypes 40 and 41. CONCLUSIONS: These findings demonstrate that NoVs and HAdVs are prevalent as virus contaminants in the karst aquifer, representing potential health risks particularly during the rainy season, in one of the most important areas used for tourism in Mexico. SIGNIFICANCE AND IMPACT OF THE STUDY: This is one of the few studies conducted in karst aquifers that provide a foundational baseline of the distribution, concentrations and diversity of NoVs and HadVs in these particular environments.

Effects of birth weight, sex and neonatal glucocorticoid overexposure on glucose-insulin dynamics in young adult horses.

In several species, adult metabolic phenotype is influenced by the intrauterine environment, often in a sex-linked manner. In horses, there is also a window of susceptibility to programming immediately after birth but whether adult glucose-insulin dynamics are altered by neonatal conditions remains unknown. Thus, this study investigated the effects of birth weight, sex and neonatal glucocorticoid overexposure on glucose-insulin dynamics of young adult horses. For the first 5 days after birth, term foals were treated with saline as a control or ACTH to raise cortisol levels to those of stressed neonates. At 1 and 2 years of age, insulin secretion and sensitivity were measured by exogenous glucose administration and hyperinsulinaemic-euglycaemic clamp, respectively. Glucose-stimulated insulin secretion was less in males than females at both ages, although there were no sex-linked differences in glucose tolerance. Insulin sensitivity was greater in females than males at 1 year but not 2 years of age. Birth weight was inversely related to the area under the glucose curve and positively correlated to insulin sensitivity at 2 years but not 1 year of age. In contrast, neonatal glucocorticoid overexposure induced by adrenocorticotropic hormone (ACTH) treatment had no effect on whole body glucose tolerance, insulin secretion or insulin sensitivity at either age, although this treatment altered insulin receptor abundance in specific skeletal muscles of the 2-year-old horses. These findings show that glucose-insulin dynamics in young adult horses are sexually dimorphic and determined by a combination of genetic and environmental factors acting during early life.

Effects of maternal dexamethasone treatment on pancreatic ß cell function in the pregnant mare and post natal foal.

REASONS FOR PERFORMING STUDY: Synthetic glucocorticoids are used to treat inflammatory conditions in horses. In other pregnant animals, glucocorticoids are given to stimulate fetal maturation with long-term metabolic consequences for the offspring if given preterm. However, their metabolic effects during equine pregnancy remain unknown. OBJECTIVE: Thus, this study investigated the metabolic effects of dexamethasone administration on pregnant pony mares and their foals after birth. STUDY DESIGN: Experimental study. METHODS: A total of 3 doses of dexamethasone (200 µg/kg bwt i.m.) were given to 6 pony mares at 48 h intervals beginning at ≈270 days of pregnancy. Control saline injections were given to 5 mares using the same protocol. After fasting overnight, pancreatic ß cell responses to exogenous glucose were measured in the mares before, during and after treatment. After birth, pancreatic ß cell responses to exogenous glucose and arginine were measured in the foals at 2 and 12 weeks. RESULTS: In mares during treatment, dexamethasone but not saline increased basal insulin concentrations and prolonged the insulin response to exogenous glucose. Basal insulin and glucose concentrations still differed significantly between the 2 groups 72 h post treatment. Dexamethasone treatment significantly reduced placental area but had little effect on foal biometry at birth or subsequently. Foal ß cell function at 2 weeks was unaffected by maternal treatment. However, by 12 weeks, pancreatic ß cell sensitivity to arginine, but not glucose, was less in foals delivered by dexamethasone- than saline-treated mares. CONCLUSIONS: Dexamethasone administration induced changes in maternal insulin-glucose dynamics, indicative of insulin resistance and had subtle longer term effects on post natal ß cell function of the foals. The programming effects of dexamethasone in horses may be mediated partially by altered maternal metabolism and placental growth.

Glucocorticoid programming of intrauterine development.

Glucocorticoids (GCs) are important environmental and maturational signals during intrauterine development. Toward term, the maturational rise in fetal glucocorticoid receptor concentrations decreases fetal growth and induces differentiation of key tissues essential for neonatal survival. When cortisol levels rise earlier in gestation as a result of suboptimal conditions for fetal growth, the switch from tissue accretion to differentiation is initiated prematurely, which alters the phenotype that develops from the genotype inherited at conception. Although this improves the chances of survival should delivery occur, it also has functional consequences for the offspring long after birth. Glucocorticoids are, therefore, also programming signals that permanently alter tissue structure and function during intrauterine development to optimize offspring fitness. However, if the postnatal environmental conditions differ from those signaled in utero, the phenotypical outcome of early-life glucocorticoid receptor overexposure may become maladaptive and lead to physiological dysfunction in the adult. This review focuses on the role of GCs in developmental programming, primarily in farm species. It examines the factors influencing GC bioavailability in utero and the effects that GCs have on the development of fetal tissues and organ systems, both at term and earlier in gestation. It also discusses the windows of susceptibility to GC overexposure in early life together with the molecular mechanisms and long-term consequences of GC programming with particular emphasis on the cardiovascular, metabolic, and endocrine phenotype of the offspring.

HORSE SPECIES SYMPOSIUM: Glucocorticoid programming of hypothalamic-pituitary-adrenal axis and metabolic function: Animal studies from mouse to horse.

Adrenal glucocorticoids, such as cortisol, are essential for normal fetal development and for maintaining homeostasis in adults. Developmental studies in humans and other animals have shown that exposure to excess glucocorticoids during critical windows of perinatal development can program permanent changes in hypothalamic-pituitary-adrenal (HPA) axis function and metabolic function, with adverse implications for the long-term health of the exposed offspring. The current review compares the programming of postnatal HPA axis function and glucose homeostasis among different species overexposed perinatally to glucocorticoids, with emphasis on the horse. The potential role of epigenetic modification of genes involved in the regulation of HPA axis and metabolic function at cellular and molecular levels is also discussed.

Early diagnosis of a Mexican variant of Papaya meleira virus (PMeV-Mx) by RT-PCR.

Papaya meleira disease was identified in Brazil in the 1980s. The disease is caused by a double-stranded RNA virus known as Papaya meleira virus (PMeV), which has also been recently reported in Mexico. However, previously reported PMeV primers failed to diagnose the Mexican form of the disease. A genomic approach was used to identify sequences of the Mexican virus isolate, referred here to as PMeV-Mx, to develop a diagnostic method. A mini cDNA library was generated using total RNA from the latex of fruits; this RNA was also sequenced using the Illumina platform. Sequences corresponding to the previously reported 669-base pair sequence for PMeV from Brazil (PMeV-Br) were identified within the PMeV-Mx genome, exhibiting 79-92% identity with PMeV-Br. In addition, a new sequence of 1154-base pairs encoding a putative RNA-dependent RNA polymerase was identified in PMeV-Mx. Primers designed against this sequence detected both virus isolates, 2 amplicons of 173 and 491 base pairs from PMeV-Br and PMeV-Mx, and shared 100 and 98% identity, respectively. PMeV-Mx was found in the latex of fruits, in seedlings, and in the leaves, flowers, petioles, and seeds of mature plants. PMeV-Mx was more abundant in the latex of fruits than in the leaves. The limit of detection of the CB38/CB39 primer pair was 1 fg and 1 pg using total RNA extracted from the latex of fruits and from seedlings, respectively. A sensitive and early diagnosis protocol was developed; this method will enable the certification of seeds and seedlings prior to transplantation to the field.

Neonatal glucocorticoid overexposure programs pituitary-adrenal function in ponies.

The present study tested the hypothesis that overexposure to endogenous glucocorticoids in neonatal life alters the reactivity of the hypothalamic-pituitary-adrenal (HPA) axis in ponies at 1 and 2 yr of age. Newborn foals received saline (0.9% NaCl, n = 8, control) or long-acting adrenocorticotropic hormone (ACTH1-24) (Depot Synacthen 0.125 mg intramuscularly twice daily, n = 9) for 5 d after birth to raise cortisol concentrations 5- to 6-fold. At 1 and 2 yr of age, HPA axis function was assessed by bolus administration of short-acting ACTH1-24 (1 µg/kg intravenous) and insulin (0.5 U/kg intravenous) to induce hypoglycemic on separate days. Arterial blood samples were taken at 5 to 30-min intervals before and after drug administration to measure plasma ACTH and/or cortisol concentrations. There were no differences in the basal plasma ACTH or cortisol concentrations or in the cortisol response to exogenous ACTH1-24 with neonatal treatment or age. At 1 and 2 yr of age, the increment in plasma ACTH but not cortisol at 60 min in response to insulin-induced hypoglycemia was greater in ponies treated neonatally with ACTH than saline (P < 0.05). Neonatal cortisol overexposure induced by neonatal ACTH treatment, therefore, alters functioning of the HPA axis in adult ponies.

Sex-associated differences in pancreatic ß cell function in healthy preweaning pony foals.

REASONS FOR PERFORMING STUDY: Pancreatic ß cells are responsive to a range of stimuli during early post natal life in healthy pony foals. However, little is known about whether these responses are sex-linked. OBJECTIVES: To determine pancreatic ß cell responses to the insulin secretagogues, glucose, arginine and tolbutamide, in fillies and colts during the first 3 months after birth. STUDY DESIGN: In vivo experiment examining sex differences in pancreatic ß cell function in foals. METHODS: Female (n = 8) and male (n = 5) pony foals were infused i.v. with glucose (0.5 g/kg bwt 40% dextrose), arginine (100 mg/kg bwt) or tolbutamide (10 or 20 mg/kg bwt) over 5 min, at 48 h intervals, to assess pancreatic ß cell function at ages 2 and 12 weeks. Blood samples (4 ml) were taken through a jugular catheter at -30, -15 and 0 min (immediately before) and 5, 15, 30, 45, 60, 90 and 120 min after glucose, arginine and tolbutamide administration for measurements of plasma glucose, α-amino-nitrogen and insulin concentrations. RESULTS: The maximum increment in plasma insulin concentration in response to glucose was significantly higher in female (395 ± 58 ng/l) than male (172 ± 37 ng/l, P<0.05) pony foals 2 weeks after birth and the area under the insulin curve was significantly greater in females at this age. At 12 weeks, the insulin increment in response to glucose was significantly greater in fillies 45 min post infusion. The ß cell responses to arginine and tolbutamide were not sex-linked at either age. CONCLUSIONS: These data show that in ponies, fillies have a greater ß cell response to glucose than colts in early post natal life. Since glucose clearance was unaffected by sex, the results suggest that fillies may be less insulin sensitive than colts shortly after birth. Innate sex differences in the secretion and action of insulin in early post natal life may influence tissue development and growth with potentially more long-term metabolic consequences.

Characterization of Rhynchosia yellow mosaic Yucatan virus, a new recombinant begomovirus associated with two fabaceous weeds in Yucatan, Mexico.

Rhynchosia minima (L.) DC. (Fabaceae) plants exhibiting bright golden mosaic symptoms were previously associated with begomovirus infection in Yucatan, México [1]. To characterize the begomovirus infecting these plants, the complete bipartite genome was cloned and sequenced. Sequence comparisons indicated that the virus was distinct from all other begomoviruses known to date, including those previously identified from symptomatic R. minima, and the name Rhynchosia yellow mosaic Yucatan virus (RhYMYuV) is proposed. Pairwise comparisons indicated that RhYMYuV DNA-A [2,597 nt, (EU021216)] and DNA-B [2,542 nt, (FJ792608)] components shared the highest nt sequence identity with Cabbage leaf curl virus (CaLCuV), 87% for component A and 71% for component B. Phylogenetic analysis indicated that both components of RhYMYuV are most closely related to other New World begomoviruses, having as closest relatives immediate outliers to the major Squash leaf curl virus (SLCV) clade. Recombination analysis of the RhYMYuV genome indicated that the DNA-A component has arisen through intermolecular recombination. R. minima plants inoculated with the monomeric clones developed a bright yellow mosaic similar to symptoms observed in naturally infected plants, confirming that the clones were infectious. Nicotiana benthamiana plants biolistically inoculated with monomeric clones developed curling and chlorosis in the newly emerging leaves. RhYMYuV was also detected in symptomatic Desmodium sect. Scorpiurus Benth. (Fabaceae) that were collected near the RhYMYuV-infected plants.