Influence of an indigenous European alder (Alnus glutinosa (L.) Gaertn) rhizobacterium (Bacillus pumilus) on the growth of alder and its rhizosphere microbial community structure in two soils

Europeanalder seedlings were inoculated with a suspension of the putative plant growthpromoting rhizobacterium (PGPR) Bacillus pumilus (CECT5105), or left non-inoculated (controls) in two different soils, and grownundercontrolled conditions. Soil A showed a thick texture, slightly acidic with ahigh mineral nitrogen content, while soil B showed a thin texture, basic andwith a lower nitrogen content. At each sampling time, over an 8-week period,shoot and root systems of the plants were measured, nodules counted, and shootand root length and surface were determined. In addition, changes in themicrobial rhizosphere structure were evaluated by the phospholipid fatty acid(PLFA) profile extracted directly from the rhizosphere soil. The increasesdetected in shoot surface were significant only in soil A, while the rootsystemwas affected in both soils. In soil A, inoculation with B.pumilus caused a perturbation that subsequently disappeared, whilethe rhizosphere community structure was seriously altered in soil B. Allbiometric parameters were enhanced to a greater extent in soil A, in which theinoculum did not alter the existing rhizosphere communities and nutrientavailability was better.     

Farm Management Effects on Rhizosphere Colonization by Native Populations of 2,4-Diacetylphloroglucinol-Producing Pseudomonas spp. and Their Contributions to Crop Health

ABSTRACT Analyses of multiple field experiments indicated that the incidence and relative abundance of root-colonizing phlD+ Pseudomonas spp. were influenced by crop rotation, tillage, organic amendments, and chemical seed treatments in subtle but reproducible ways. In no-till corn plots, 2-year rotations with soybean resulted in plants with approximately twofold fewer phlD+ pseudomonads per gram of root, but 3-year rotations with oat and hay led to population increases of the same magnitude. Interestingly, tillage inverted these observed effects of cropping sequence in two consecutive growing seasons, indicating a complex but reproducible interaction between rotation and tillage on the rhizosphere abundance of 2,4-diacetlyphloroglucinol (DAPG) producers. Amending conventionally managed sweet corn plots with dairy manure compost improved plant health and also increased the incidence of root colonization when compared with nonamended plots. Soil pH was negatively correlated to rhizosphere abundance of phlD+ pseudomonads in no-till and nonamended soils, with the exception of the continuous corn treatments. Chemical seed treatments intended to control fungal pathogens and insect pests on corn also led to more abundant populations of phlD in different tilled soils. However, increased root disease severity generally was associated with elevated levels of root colonization by phlD+ pseudomonads in no-till plots. Interestingly, within a cropping sequence treatment, correlations between the relative abundance of phlD and crop stand or yield were generally positive on corn, and the strength of those correlations was greater in plots experiencing more root disease pressure. In contrast, such correlations were generally negative in soybean, a difference that may be partially explained by difference in application of N fertilizers and soil pH. Our findings indicate that farming practices can alter the relative abundance and incidence of phlD+ pseudomonads in the rhizosphere and that practices that reduce root disease severity (i.e., rotation, tillage, and chemical seed treatment) are not universally linked to increased root colonization by DAPG-producers.     

SSR markers for marker assisted selection of root-knot nematode (<Emphasis Type="Italic">Meloidogyne incognita</Emphasis>) resistant plants in cotton (<Emphasis Type="Italic">Gossypium hirsutum</Emphasis> L)

Cotton (Gossypium hirsutum L) cultivars highly resistant to the southern root-knot nematode (RKN) [Meloidogyne incognita (Kofoid and White) Chitwood] are not available. Resistant germplasm lines are available; however, the difficulty of selecting true breeding lines has hindered applied breeding and no highly resistant cultivars are available to growers. Recently, molecular markers on chromosomes 11 and 14 have been associated with RKN resistance, thus opening the way for marker assisted selection (MAS) in applied breeding. Our study aimed to determine the utility of these markers for MAS. Cross one was RKN resistant germplasm M240 RNR × the susceptible cultivar, FM966 and is representative of the initial cross a breeder would make to develop a RKN resistant cultivar. Cross two consists of Clevewilt 6 × Mexico Wild (PI563649), which are the two lines originally used to develop the first highly RKN resistant germplasm. Mexico Wild is photoperiodic. We phenotyped the F₂ of cross one for gall index and number of RKN eggs per plant and genotyped each plant for CIR 316 (chromosome 11) and BNL 3661 (chromosome 14). From this, we verified that MAS was effective, and the QTL on chromosome 14 was primarily associated with a dominant RKN resistance gene affecting reproduction. In the first F₂ population of cross two, we used MAS to identify 11 plants homozygous for the markers on chromosomes 11 and 14, and which also flowered in long days. Progeny of these 11 plants were phenotyped for RKN gall index and egg number and confirmed as RKN highly resistant plants. Generally about 7–10 generations of RKN phenotyping and progeny testing were required to develop the original RKN highly resistant germplasms. Our results show that commercial breeders should be able to use the markers in MAS to rapidly develop RKN resistant cultivars.     

Increased incidence of DNA amplification in follicular than in uterine and blood samples indicates possible tropism of Neospora caninum to the ovarian follicle

This study evaluated the presence of Neospora caninum in ovarian follicle aspirates and uterine flushes obtained from N. caninum seropositive dairy cows. Ninety-two cows that aborted within the previous 90 days were sampled to determine the presence of antibodies against N. caninum. Thirteen seropositive cows were chosen for collection of blood leukocytes, uterine flushes (UF; n=12) and follicular aspirates (OPU; n=13). Samples were centrifuged and the cellular sediment from the follicular fluid, uterine flushes and blood leukocytes were used for DNA extraction and PCR. Follicular aspirates had the highest frequency of DNA amplification for N. caninum (p<0.05, 92.3%; 12/13). Whereas uterine (4/12) and blood leukocyte (5/13) samples had similar (p>0.05) rate of positive results. Nonetheless, there was no agreement between blood leukocytes and follicular samples taken from the same animal (Cohen Kappa=-0.16). Similarly, blood leukocytes and uterine results had moderate agreement between them (Cohen Kappa=0.47). This study indicates that N. caninum is present in the ovarian follicle and uterus of seropositive cows, suggesting a possible risk of neosporosis transmission between females during oocyte and embryo collection and transfer. Hence, precautions should be taken to minimize the risk of N. caninum transmission. Furthermore, the high incidence of positive results in follicle samples, that exceeded that of their paired blood leukocytes, suggests a possible tropism of N. caninum for the ovarian follicle.     

Expression of the μ Opioid Receptor and Effects of the Opioid Antagonist Naloxone on In Vitro Maturation of Oocytes Recovered from Anoestrous Bitches

The μ-opioid receptor (MOR) is expressed in bovine, human, equine and canine oocytes, and in seasonal breeders, it is expressed with higher intensity during the anoestrous phase. Supplementation of in vitro maturation (IVM) medium with opioid agents, agonists or antagonists, was shown to affect oocyte maturation in several species such as rat, bovine and equine. This study reports the effects of supplementing IVM medium with naloxone (Nx), an opioid antagonist, on nuclear and cytoplasmic maturation rate of oocytes recovered from anoestrous bitches. Cytoplasmic maturation was examined in terms of mitochondrial (mt) distribution. In order to confirm the receptor-mediated action of Nx, in oocytes of anoestrous bitches, MOR expression was analyzed by Western blot. Cumulus–oocyte complexes, recovered from the ovaries of bitches in anoestrous, were cultured in vitro and Nx was added at the concentrations of 1 × 10⁻⁶, 1 × 10⁻⁸ and 1 × 10⁻¹⁰  m . The rate of oocytes resuming meiosis after culture in presence of 1 × 10⁻⁶  m Nx (29%) was significantly higher than that of oocytes of control group (12%; p < 0.05). However, treatment with Nx did not affect mt distribution pattern. In denuded oocytes and in corresponding cumulus cells, a doublet of 65 and 50 kDa was observed. We conclude that, in oocytes of anoestrous bitches, MOR is expressed and Nx significantly improves nuclear maturation rate. Further studies should be performed to elucidate the expression of other opioid receptors, such as δ and κ, and possible interactive effects of their antagonists on canine oocyte maturation.     

Serological evidence of Ehrlichia spp. exposure in cats from northeastern Spain

There is little information about Ehrlichia canis as an infectious agent in cats. In order to estimate the prevalence of antibodies to E. canis in the feline population, 235 cat sera were analysed by indirect fluorescent-antibody test. With the objective to determine some risk factors associated with seropositivity, serum samples were divided into two groups: urban stray cats and pet cats. The seroprevalence detected was 17.9%. Most positive sera (83.3%) showed low antibody titres (<1:80). Seropositivity was very similar when comparing the two groups of animals: 17.4% in urban stray cats and 18.4% in pet cats. Results revealed that cats are exposed to Ehrlichia spp. infection, as the low antibody titres detected and the serological cross-reactivity between Ehrlichia species do not allow us to confirm E. canis exposure.     

Second Oestrus Synchronization and Precocious Embryo Viability after Puberty Induction in Gilts by the Use of Gonadotrophin Treatment

The use of exogenous gonadotrophins in puberty inducement and ovulation synchronization is a technique that has a positive influence on the management of swine. The purpose of this study was to verify the effects of a second gonadotrophin treatment [equine chorionic gonadotrophin (eCG) and luteinizing hormone (LH), intramuscularly (i.m.)] upon the second oestrus synchronization and fertility in gilts. Seventy-one NAIMA (Pen Ar Lan) gilts had their first oestrus (puberty inducement) induced by a hormonal treatment (eCG and LH). Then, they were randomly distributed into two treatments, with (T1) and without (C) gonadotrophin treatment at the second oestrus. The animals were fed with a single ration (16% of crude protein and 3286.73 kcal ME/kg), and timed artificial insemination performed at the second oestrus. Gilts were slaughtered for embryo recovery and ovary examination about 5 days after insemination. There was no evidence of a difference in the percentage of the second oestrus expression (T1 - 90.90% and C - 86.84%), the duration of the oestrus cycle (T1 - 19.62 +/- 0.82 days and C - 19.67 +/- 4.14 days), the percentage of follicular cysts (T1 - 15.15% and C - 18.42%) and number of ovulations (T1 - 14.60 +/- 5.7 and C - 13.23 +/- 4.8) between treatments (p > 0.05). However, the hormonal treatment (T1) showed minor oestrus dispersion and embryo viability (T1 - 8.4 +/- 5.6 and C - 11.2 +/- 4.6) (p < 0.05). These results indicate that the better synchronization and expression of the second oestrus when using gonadotrophins (eCG and LH) is followed by a lower embryo viability, which is probably the consequence of the heterogeneous follicle recruitment during the injection of eCG.     

Time course of endocrine changes in the hypophysis-gonad axis induced by hypobaric hypoxia in male rats

Chronic hypobaric hypoxia (CHH) induces a decrease in sperm output and spermatogenesis in male rats. The mechanisms that underlie these changes in testicular function are unknown and could involve changes in the hypophysis-gonad axis. We have tested the hypothesis that changes take place in the endocrine status (FSH, follicle stimulating hormone; LH, luteinizing hormone; testosterone) of rats subjected to CHH. Male Wistar rats were maintained under normobaric or hypobaric conditions (428 torr, 4,600 m). On days 0, 5, 15 and 30 post-exposure, 12 rats were anesthetized, their body weights were measured and blood samples were collected. The testicles were fixed in 4% formaldehyde and processed for histological analysis. In this time course, the FSH levels rose by day 5 post-exposure. On subsequent days, the FSH levels decreased in rats subjected to CHH with a tendency to remain higher than the normoxic group. The LH plasma levels decreased in rats exposed to CHH. Consistent with the decrease in LH levels, the plasma testosterone level decreased significantly after 30 days of CHH exposure. Integrated analysis of hormonal changes in rats subjected to CHH and the body dehydration that occurs in HH allows us to conclude that the effects of CHH on spermatogenesis may be partially related to changes in the hypophysis-gonad hormonal axis.