Expression of MHC-I and -II in Uterine Tissue from Early Pregnant Bitches

The aim of the study was to investigate the expression of major histocompatibility complex (MHC)-I and -II in uterine tissues from pregnant and non-pregnant bitches, taken at different time periods after mating. The pregnant bitches were ovariohysterectomized during the pre-implantation (group 1, n = 4), implantation (group 2, n = 7) and placentation stage (group 3, n = 7). Non-pregnant animals in diestrus served as controls (group 4, n = 7). The expression of MHC- I and -II in salpinx, apex, middle horn, corpus uteri and at implantation sites was investigated by immunohistochemistry as well as qualitative and quantitative RT-PCR; MHC-I mRNA was detected in all tissues and with quantitative RT-PCR, and no significant changes were detected until placentation. Immunohistologically, at the apex and corpus site, the average number of MHC-II positive cells increased from the pre-implantation to the post-implantation stage (apex: 1.54 ± 1.21 to 3.82 ± 2.93; corpus: 1.62 ± 1.9 to 5.04 ± 4.95; p < 0.05). The greatest numbers of MHC-II positive cells were observed at placentation sites (6.64 ± 5.9). In parallel, a marked increase in the relative mRNA expression of MHC-II in uterine tissues was assessed from the pre-implantation to the placentation stage (relative to Glycerinaldehyd-3-phosphate-Dehydrogenase (GAPDH): 6.9 ± 9.5, 8.4 ± 5.8, p > 0.05). Immunohistologically, in the salpinx, significantly greater numbers of MHC-II positive cells were found in the tissues of pregnant animals than in the control group (p < 0.05). It is proposed that the increase in MHC-II is pregnancy-related, even though the impact on maintenance of canine pregnancy is still unclear.     

Volatile organic compounds in watermelon (Citrullus lanatus) grafted onto 21 local and two commercial bottle gourd (Lagenaria siceraria) rootstocks

Summary

Volatile organic compounds (VOCs) are responsible for flavour development in watermelon (Citrullus lanatus). The objective of this study was to investigate the rootstock effects of 21 local bottle gourd [Lagenaria siceraria (Molina) Standl.] accessions collected from the Mediterranean region (Turkey) on the concentrations of VOCs in watermelon over 2 years. The watermelon cultivar ‘Crimson Tide’ was used as the scion and two commercial bottle gourd rootstocks were also used for comparison. The VOCs in each rootstock-scion combination were analysed using static headspace (SHS) and gas chromatography-mass spectrometry (GC-MS). A total of 24 VOCs were identified in watermelon fruit, including 11 aldehydes, seven alcohols, four ketones, one heterocyclic compound, and one acid. Acetaldehyde (25.9 – 39.5% of the total peak areas), propanal (5.0 – 8.3%), 2-propanone (2.5 – 4.9%), pentanal (2.1 – 3.6%), hexenal (14.5 – 22.6%), (E)-2-hexenal (0.4 – 2.3%), 6-methyl-5-hepten-2-one (10.0 – 19.05%), nonanal (1.6 – 4.4%), (Z)-6-nonenal (4.0 – 9.8%), (E)-2-nonenal (0.6 – 2.0%), (E,Z)-2,6-nonadienal (2.1 – 3.7%), (Z)-6-nonenol (1.0 – 3.2%), and (E,Z)-3,6-nonadienol (1.0 – 3.7%) were the most abundant VOCs found in all watermelon scions. There was no significant effect (P ≥ 0.05) of rootstock on the concentrations of acetaldehyde, propanal, pentanal, heptanal, ethanol, 1-pentanol, 2-propanone, 2-pentyl furan, or acetic acid. Propanal, 2-propanone, and acetic acid were identified as VOCs in watermelon for the first time. Hexanal, (Z)-6-nonanal, and 6-methyl-5-hepten-2-one concentrations were strongly affected by rootstock. The results showed that the local bottle gourd accessions 07-06 and 31-09, in particular, were the most promising genotypes in terms of producing favourable VOCs. Except for the local bottle gourd accessions 33-35, 01-17, and 07-04, the other 18 gourds accessions had high potential as rootstocks for breeding programmes in watermelon with regard to desirable VOC concentrations. Rootstock 31-09 should be considered for commercial production in terms of improved disease resistance, fruit yield, and fruit quality.     

Screening mycorrhiza species for plant growth,P and Zn uptake in pepper seedling grown under greenhouse conditions

We screened the mycorrhizal species for an inoculum protocol would green pepper seedling production and compensate for nutrient deficiency. Three greenhouse studies (on three successive years) were conducted under nursery conditions using five arbuscular mycorrhizal (AM) fungi and their mixture. The AM fungi, Glomus mosseae, G. clarum, G. caledonium, G. intraradices and G. etunicatum propagated on maize roots were used. The AM fungi were inoculated to seed stages and half of the seed stages inoculated plants were re-inoculated at the seedling stages. In Experiment I, plants were harvested once and in Experiments II and III, plants were harvested twice at different developmental stages.     

Vascular Endothelial (VEGF) and Epithelial Growth Factor (EGF) as Well as Platelet‐Activating Factor (PAF) and Receptors are Expressed in the Early Pregnant Canine Uterus

The aim of this study was to investigate the course of expression of platelet‐activating factor (PAF), PAF‐receptor (PAF‐R), epidermal growth factor (EGF), EGF‐R, vascular endothelial growth factor (VEGF), VEGF‐R1 and VEGF‐R2 in uterine tissue during canine pregnancy. For this purpose, 20 bitches were ovariohysterectomized at days 10–12 (n = 10), 18–25 (n = 5) and 28–45 (n = 5) days after mating, respectively. The pre‐implantation group was proven pregnant by embryo flushing of the uterus after the operation, the others by sonography. Five embryo negative, that is, non‐pregnant, bitches in diestrus (day 10–12) served as controls. Tissue samples from the uterus (placentation sites and horn width, respectively) were excised and snap‐frozen in liquid nitrogen after embedding in Tissue Tec^®. Extraction of mRNA for RT‐PCR was performed with Tri‐Reagent. In the embryos, mRNA from all factors except VEGF was detected. In the course of pregnancy, significantly higher expression of PAF and PAFR as well as VEGF and VEGFR2 during the pre‐implantation stage than in all other stages and a strong upregulation of EGF during implantation were characteristic. The course of EGF was in diametrical opposition to the course of the receptor. These results point towards an increased demand for VEGF, EGF and PAF during the earliest stages of canine pregnancy.     

Expression and Activity of Matrix Metalloproteinases in the Uterus of Bitches After Spontaneous and Induced Abortion

Aim of this study was to determine the intrauterine activity of matrix metalloproteinases (MMP)‐2 and ‐9 after cessation of the local effect of progesterone. For this purpose, pregnancy was terminated in 10 bitches at mid‐gestation with the progesterone receptor antagonist aglepristone (10 mg/kg body weight, sc, Alizine^®; Virbac, France) at two subsequent days (group IRA = induced resorption/abortion). The IRA group was divided into two subgroups (Group I, n = 5, days 25–35 of pregnancy; group II, n = 5, days 36–45). Five further bitches were introduced with beginning abortion (group SRA = spontaneous resorption/abortion). Seven healthy bitches between day 25 and 45 of gestation served as controls. After ovariohysterectomy at the end of abortion and between days 25 and 45 of gestation, respectively, the distribution and activity of collagenases were investigated by immunohistochemistry and gelatin zymography. At placental sites, MMP‐2 activity in the endometrium was significantly lower in IRA groups than in the SRA group (33.7 ± 11.8% and 39.3 ± 5.4% vs 52.2 ± 10.2%, p < 0.05); however, MMP‐2 expression was lowest in the control group (control: 21.4 ± 6.3%; p < 0.01) and similarly in the myometrium (controls: 13.1 ± 2.5%; p < 0.05). MMP‐9 activity was also lower in the endometrium and myometrium of the control group in comparison to SRA and IRA groups (11.8 ± 3.2%; p < 0.01 and 28.4 ± 32.8%; p < 0.05). At interplacental sites, the amount of active collagenases in the myometrium was significantly lower in the control group. It is concluded that the blockade of the biological progesterone effect was associated with an increase in activity of both collagenases.     

RESPONSES OF GRAFTED WATERMELON ONTO DIFFERENT GOURD SPECIES TO SALINITY STRESS

The watermelon cultivar ‘Crimson Tide’ was grafted onto three different rootstocks and grown under saline conditions to investigate effects of salinity on grafted and non-grafted watermelon. One Cucurbita maxima and 2 Lagenarai siceraria landraces (Skp and Brecik) were used as rootstock. Plants were irrigated with two different saline solutions [0.5 (control) and 8.0 dS m^?1] by two days interval at the first 15 days of experiment and one day interval at the last 15 days of experiment. The experimental design was randomized block. Each treatment was replicated three times with three plants. Grafted plants had higher plant growth parameter than non-grafted plant under saline conditions. Reduction in shoot dry weight was 41% in non-grafted plants while it was varied from 22% to 0.8% in grafted plants under saline conditions. Accumulation of sodium (Na⁺) was higher in non-grafted plants than grafted one. Calcium (Ca⁺⁺) and magnesium (Mg⁺⁺) concentration were higher in all grafted plants than non-grafted plants. Non-grafted plants had higher K⁺ concentration than other treatments under saline conditions. Ratios of Ca⁺⁺/Na⁺, K⁺/Na⁺ and Mg⁺⁺/Na⁺ were significantly affected by salt treatments and positively correlated with plant growth parameters. The ratios were lower in non-grafted plants than grafted plants under saline conditions.     

Rootstock resistance to Fusarium wilt and effect on watermelon fruit yield and quality

The potential of grafted watermelon for resistance toFusarium oxysporum f.sp.niveum on some Curcurbitaceae,Lagenaria, Luffa, Benincasa and commercial rootstocks was evaluated. Effects of grafting on yield and quality of diseased plants were evaluated. All grafted plants and rootstocks were resistant to the three known races (0, 1, and 2) ofF. oxysporum f.sp.niveum except watermelon cv. ‘Crimson Tide’, which was susceptible to race 2. Fruit yield was positively (21–112%) affected byLagenaria rootstocks but negatively affected (200–267%) byCucurbita rootstocks when compared with the control. While only minor differences in fruit quality were determined in control and grafted plants onLagenaria rootstocks, the quality parameters for watermelon grafted ontoCucurbita rootstocks were lower than in the control. The reasons for low yield and quality might be due to an incompatibility betweenCucurbita rootstocks and watermelon. These results showed that rootstock influence on disease resistance as well as yield and quality of scion fruit is important in determining the potential use of grafting applications in watermelon. http://www.phytoparasitica.org posting Feb. 2, 2003.     

A Screening for the Occurrence of Autoreactive Antibodies in Sera of Pregnant and Non-pregnant Bitches

Sera of healthy pregnant (group I, n = 11) and non-pregnant (group II, n = 11) bitches were screened for autoantibodies (AAb). In both groups, blood samples were drawn every fifth day between days 5 and 55 after mating. Serum was analysed via indirect immunofluorescence (IIF) with the Canine ANA HEp-2 Screening Kit. In all animals, anticytoplasmic AAb were detected. Utilizing primate-heart substrate slides AAb against contractile proteins of the cytoplasm could be observed. The predominating fluorescence pattern in pregnant animals resembled above all desmin, which was proven via Western blot. The sera were then pre-incubated with tropomyosin, actin, vimentin, vinculin and keratin solutions, and assessed on HEp-2 slides and on human and canine fibroblasts as well. The latter substrate was used to verify whether the detected Ab were in fact AAb. Utilizing tropomyosin, revealed elimination of the cytoplasmic fluorescences on all three substrates. It is therefore assumed, that in sera of healthy dogs, AAb against contractile structure proteins of the cytoplasm are present regularly. The majority of pregnant bitches presented with higher end-point titres (EPT), than to be found in non-pregnant dogs. AAb against desmin played the key role in those patterns. In addition, sera were screened for thyroid specific AAb, namely thyroglobulin, thyroid peroxidase (TPO), T3 and T4, and for AAb against insulin by ELISA or Western blot (TPO). Only in two of the pregnant bitches a weak positive reaction (1:100) for T3-AAb was detected.