Advantages of real-time PCR assay for diagnosis and monitoring of canine leishmaniosis

The aim of the present study is to highlight the advantages of real-time quantitative PCR intended to aid in the diagnosis and monitoring of canine leishmaniosis. Diagnosis of canine leishmaniosis is extremely challenging, especially in endemic areas, due to the diverse and non-specific clinical manifestations, and due to the high seroprevalence rate in sub-clinical dogs. Veterinarian clinicians are usually confronted with cases that are compatible with the disease, and with several diagnostic tests, sometimes with contradictory results. We have developed a new TaqMan assay, targeting the kinetoplast, applied to 44 samples of bone marrow aspirate or peripheral blood. The dynamic range of detection of Leishmania DNA was established in 7 logs and the limit of detection is 0.001 parasites in the PCR reaction. At the time of diagnosis parasitemia ranges from less than 1 to 10(7)parasites/ml. The ability to quantify the parasite burden allowed: (i) to elucidate the status of positive dogs by conventional PCR, although larger studies are necessary to clarify the dividing line between infection and disease, (ii) to estimate the kinetics of the parasite load and the different response to the treatment in a follow-up and (iii) to validate blood as less invasive sample for qPCR. The continuous data provided by real-time qPCR could solve the dilemma for the clinician managing cases of canine leishmaniosis by differentiating between Leishmania-infected dogs or dogs with active disease of leishmaniosis.     

Effect of changes in ionized calcium concentration in arterial blood and metabolic acidosis on the arterial partial pressure of oxygen in dogs

OBJECTIVE: To evaluate the effects of metabolic acidosis and changes in ionized calcium (Ca2+) concentration on PaO2 in dogs. ANIMALS: 33 anesthetized dogs receiving assisted ventilation. PROCEDURE: Normal acid-base status was maintained in 8 dogs (group I), and metabolic acidosis was induced in 25 dogs. For 60 minutes, normocalcemia was maintained in group I and 10 other dogs (group II), and 10 dogs were allowed to become hypercalcemic (group III); hypocalcemia was then induced in groups I and II. Groups II and IV (5 dogs) were treated identically except that, at 90 minutes, the latter underwent parathyroidectomy. At intervals, variables including PaO2, Ca2+ concentration, arterial blood pH (pHa), and systolic blood pressure were assessed. RESULTS: In group II, PaO2 increased from baseline value (96 +/- 2 mm Hg) within 10 minutes (pHa, 7.33 +/- 0.001); at 60 minutes (pHa, 7.21 +/- 0.02), PaO2 was 108 +/- 2 mm Hg. For the same pHa decrease, the PaO2 increase was less in group III. In group I, hypocalcemia caused PaO2 to progressively increase (from 95 +/- 2 mm Hg to 104 +/- 3 mm Hg), which correlated (r = -0.66) significantly with a decrease in systolic blood pressure (from 156 +/- 9 mm Hg to 118 +/- 10 mm Hg). Parathyroidectomy did not alter PaO2 values. CONCLUSIONS AND CLINICAL RELEVANCE: Induction of hypocalcemia and metabolic acidosis each increased PaO2 in anesthetized dogs, whereas acidosis-induced hypercalcemia attenuated that increase. In anesthetized dogs, development of metabolic acidosis or hypocalcemia is likely to affect ventilatory control.     

Intravaginal progesterone devices in synchronization protocols for artificial insemination in beef heifers

Two experiments were designed to investigate the administration of intravaginal progesterone in protocols for oestrus and ovulation synchronization in beef heifers. In Experiment 1, cyclic Black Angus heifers (n = 20) received an Ovsynch protocol and were randomly assigned to receive (CIDR‐Ovsynch) or not (Ovsynch) a progesterone device between Days 0 and 7. Treatment with a controlled internal drug release (CIDR) device significantly increased the size of the dominant follicle prior to ovulation (12.8 ± 0.4 CIDR‐Ovsynch vs 11.4 ± 0.4 Ovsynch) (p < 0.02). Plasma progesterone concentrations throughout the experiment were affected by the interaction between group and day effects (p < 0.004). In Experiment 2, cyclic Polled Hereford heifers (n = 382) were randomly assigned to one of the six treatment groups (3 × 2 factorial design) to receive a CIDR, a used bovine intravaginal device (DIB), or a medroxiprogesterone acetate (MAP) sponge and GnRH analogues (lecirelin or buserelin). All heifers received oestradiol benzoate plus one of the devices on Day 0 and PGF on Day 7 pm (device withdrawal). Heifers were detected in oestrus 36 h after PGF and inseminated 8–12 h later, while the remainder received GnRH 48 h after PGF and were inseminated on Day 10 (60 h). The number of heifers detected in oestrus on Day 8 and conception rate to AI on Day 9 were higher (p < 0.01) in the used‐DIB than in the CIDR or MAP groups, while the opposite occurred with the pregnancy rate to FTAI on Day 10 (p < 0.01). There was no effect of progesterone source, GnRH analogue or their interaction on overall pregnancy rates (64.9%). Progesterone treatment of heifers during an Ovsynch protocol resulted in a larger pre‐ovulatory follicle in beef heifers. Progesterone content of intravaginal devices in synchronization protocols is important for the timing of AI, as the use of low‐progesterone devices can shorten the interval to oestrus.     

Bovine rotavirus strains circulating in beef and dairy herds in Argentina from 2004 to 2010

Bovine Group A Rotavirus (RVA) is one of the main causes of neonatal calf diarrhea worldwide. The present study reports the genotyping of bovine RVA strains circulating in Argentinean cattle from 2004 to 2010. Additionally, a new set of typing primers was designed and tested to differentiate between G8 and G6 (lineage III and IV) RVA strains. Bovine RVA was detected in 30% (435/1462) of the tested samples, corresponding to 49% (207/423) of the studied outbreaks with a similar detection rates in beef (53%; 67/127) and dairy herds (52%; 65/126). The RVA strains circulating in Argentinean cattle belonged to the common bovine genotypes G6 (lineages III and IV), G8, G10, P[5] and P[11]. A different RVA G/P-genotype distribution was found between the exploitation types, with the combination G6(IV)P[5] being by fare the most prevalent RVA strain in beef herds (58%), whereas a more even distribution of G6(III)P[11] (15%), G10P[11] (17%), G6(IV)P[5] (14%), and G6(IV)P[11] (6%) RVA strains was detected in dairy herds. G8 RVA strains were found in two dairy farms in calves co-infected with G8+G6(III)P[11]. A high percentage of co-infections and co-circulation of RVA strains with different genotypes during the same outbreak were registered in both exploitation types (20% of the outbreaks from beef herds and 23% from dairy herds), indicating a potential environment for reassortment. This finding is significant because G10P[11] and G6(III)P[11] strains may possess zoonotic potential. Continuous surveillance of the RVA strains circulating in livestock provides valuable information for a better understanding of rotavirus ecology and epidemiology.     

Ploidy levels of Dioscorea alata L. germplasm determined by flow cytometry

Dioscorea alata L. is a highly important crop, widely distributed in the humid and semi-humid tropics. Flow cytometry was used to determine the ploidy levels of 74 D. alata genotypes collected mainly from West African countries. Sixty three of the genotypes were found to be tetraploid, one was hexaploid and ten were octoploid. The high percentage of tetraploids together with the small percentage of hexaploid individuals and the absence of diploid individuals gives us some more clues on the possible origin of these species. No association between ploidy level and place of cultivation was found for the tested material. The obtained results represent important knowledge for enhancing the breeding methodologies and optimize germplasm management of this species. It also offers further insights to the phylogeny and evolution of Dioscorea species.     

Theileria buffeli infection of a Michigan cow confirmed by small subunit ribosomal RNA gene analysis

Theileria buffeli, generally a benign parasite of cattle, has been reported in animals from Texas, Missouri and North Carolina. To date, there have been no reports of the parasite in cattle residing in northern portions of the US. An 8-year-old cow (Maine Anjou x Angus cross-bred) in Michigan presented with hemoglobinuria and a packed cell volume of 9. Blood films stained with Giemsa showed numerous intraerythrocytic parasites morphologically consistent with T. buffeli. The parasite was confirmed to be T. buffeli by SSU rRNA gene sequence analysis (SSU rRNA sequence, Type A). This represents the first report of this parasite in an animal in Michigan.     

Clues to wood quality and production from analyzing ring width and density variabilities of fertilized Pinus taeda trees

Tree growth and wood density are influenced by forest management. Nevertheless, few studies have evaluated their variability responses to fertilizer treatments at inter-annual, inter-tree and stand-production levels. Therefore, the annual ring width (RW) and density (RD) of sixty 17-year-old-Pinus taeda trees fertilized with six doses of composted pulp-mill sludge (CPMS) were analyzed. Ten trees for each treatment were felled and from which wood discs were taken at different trunk heights. The annual RW and RD were provided by X-ray microdensitometry, synchronized and the trunk basic specific gravity (SG_B) and biomass calculated. The effects of CPMS treatments were explored using interaction of variables RW and RD with cambial age, diameter, trunk SG_B and biomass production. Trees treated with CPMS grow faster, increasing their biomass (up to 108%), presenting lower wood density values (significant up to the 6th year) and reaching the mature wood later than untreated trees. Furthermore, the potential use of RW and RD in allometric equations showed good accuracy to predict trunk SG_B and biomass. Altogether, our results indicated that ring width and density revealed the impacts of fertilization treatment on wood quality and production. Our study also provides useful information for forest managers on the fertilization monitoring process.

     

Rhodotorulic acid enhances root colonization of tomato plants by arbuscular mycorrhizal (AM) fungi due to its stimulatory effect on the pre-symbiotic stages of the AM fungi

Exudates of Rhodotorula mucilaginosa, a yeast commonly found in the rhizosphere, increased hyphal length of the arbuscular mycorrhizal (AM) fungi Gigaspora rosea and Gigaspora margarita. Rhodotorulic acid (RA), a siderophore compound obtained from R. mucilaginosa exudates, increased hyphal length and branching. Thus, the increase in the number of entry points and the higher AM root colonization of tomato plants in the presence of RA can at least partially be explained by the positive effect of RA on the pre-symbiotic stages of the AM fungi.     

Characterization and growth conditions of Vibrio parahaemolyticus strains with different virulence degrees that cause acute hepatopancreatic necrosis disease in Litopenaeus vannamei

The phenotypic characteristics and growth kinetics at several temperatures, salinities, and pH values of three Vibrio parahaemolyticus (Vp) strains with different virulence and one nonpathogenic strain were evaluated. Independent of the virulence of the strain, a high metabolic diversity was found, which yielded different colored phenotypes on the CHROMagar? Vibrio. All strains were resistant to ampicillin and carbenicillin, and Vp AHPND+ organisms were the most sensitive to enrofloxacin. The exponential growth of Vp strains started at 1–2 hr of incubation, although no relationship was observed between the bacterial density and degree of virulence. Moreover, the growth of the most virulent strain was independent of the nutrients in the incubation media during the initial hour postinoculation. No strain grew at 4°C in 0% NaCl and pH 4, but only Vp AHPND+ grew at 44°C. For all strains, the lag phase was proportional to the NaCl concentration, and the growth was better at pH 8–9. However, the Vp AHPND? strain displayed a greater variability, was more sensitive to extreme conditions, and showed a lag phase of 9 hr independent of the pH.     

The effects of sex and age on phytohaemagglutinin skin-testing of deer

AIM: To determine if there are sex- or age-related differences in the increase in skinfold thickness in response to the mitogen phytohaemagglutinin (PHA) in red deer. METHODS: One dose of 250 mug PHA was injected intradermally in the right side of the neck, and phosphate buffered saline (PBS) was injected at a second site as a control, in 110 (51 males and 59 females) captive Iberian red deer (Cervus elaphus hispanicus), ranging in age from 21 months to > or =5 years. Skinfold thicknesses were measured immediately before and 72 h following injection. RESULTS: There was a significant effect of gender on the average increase in skinfold thickness; males had greater increases (8.8 (SEM 0.57) mm) than females (4.23 (SEM 0.39) mm) after correcting for other confounding variables. No age-related differences were evident, but differences between sexes were more marked with increasing age. CONCLUSIONS AND CLINICAL RELEVANCE: Effects of gender, probably due to differences in energetic and reproductive constraints in red deer, should be taken into account when interpreting skinfold-test data, both in ecology and in the control of tuberculosis (Tb). Males tend to have a thicker skin than females, so skinfold increase relative to the thickness of the skin, rather than skinfold increase per se, should be used as a more appropriate measure of skinfold increase. This may also have clinical relevance in the interpretation of tuberculin skin testing.