Adherence to various host cell lines of Mycoplasma bovis strains differing in pathogenic and cultural features

Mycoplasma bovis is known to be responsible for pneumonia and arthritis in calves, as well as mastitis in dairy cows. Despite clear evidence of its pathogenic potential, little is known about mechanisms of cytadherence and the molecular factors involved. The purpose of this work was to compare adherence rates of M. bovis field strains to different host cell lines and study the effects of cloning and sub-culturing M. bovis strains on their adherence properties. Eighteen metabolically labeled M. bovis strains isolated from different pathological backgrounds were examined in adherence trials using four different host cell lines, i.e. embryonic bovine lung (EBL), embryonic bovine trachea (EBTr), Madin Darby bovine kidney (MDBK) and rabbit kidney (RK) cells. Although large interstrain variations in adherence rates (3.4-19.1%) were measured they could not be correlated to the pathological background (pneumonia, arthritis or mastitis). Adherence rates to the fibroblast cell line (EBTr) were significantly lower than those to the three epithelial cell lines (EBL, MDBK and RK). The only non-pathogenic strain (221/89) exhibited lower adherence rates than three isolates from clinical mastitis. Interestingly, adherence rates were significantly reduced after in vitro passaging. In contrast, no effect of single cloning of strains on adherence was observed. There was no general correlation between expression of variable surface proteins (Vsps) as monitored by immunoblotting and adherence rates, although alterations in Vsp expression profiles were seen as a consequence of passaging. As there is probably a large number of adhesins, variable and non-variable, on the surface of M. bovis cells the issue is very complex, and the most active components have yet to be identified.     

Neospora caninum and Waddlia chondrophila strain 2032/99 in a septic stillborn calf

Characteristics of an intracellularly growing micro-organism isolated from an aborted bovine foetus are described. The organism replicated within cytoplasmic vacuoles, was resistant to penicillin and exhibited structural characteristics compatible with Waddlia chondrophila. An ELISA specific for Chlamydia spp., immunofluorescence tests using antibodies directed against Chlamydia spp. or Simkania negevensis, and PCR using Chlamydia-specific primers showed that the agent was distinct from Chlamydiae or S. negevensis. Determination of 16S and partial 23S ribosomal RNA gene sequences in combination with the PCR results and the morphological, antigenic and developmental characteristics provided evidence that the isolate 2032/99 can be classified as W. chondrophila or a closely related organism.     

Vitamin C Requirements of the Angelfish Pterophylum scalare

Abstract.— Despite a large economical value of the ornamental fish trade, little information is available on the dietary ascorbic acid requirements of these species. Ascorbic acid requirements of fishes of the cichlid family appear to vary widely. Juvenile angelfish, a widely produced ornamental cichlid, were maintained on diets containing graded levels of ascorbyl monophosphate. Liver ascorbic acid concentrations after 96 d of feeding were significantly reduced in groups receiving 120 mg or less ascorbic acid equivalents/kg diet. However, no differences in growth or mortality between groups were found, and no external signs of ascorbic acid deficiency were observed, indicating a high resistance of this species against prolonged ascorbic acid deficiency. Based on the long possible life span of angelfish in the aquarium, we proposed a conservative dietary ascorbic acid requirement of 360 mg/ kg diet, necessary to maintain maximum tissue storage of this vitamin.     

Effects of Temperature and Photoperiod on Reproduction of Female Yellow Perch Perca flavescens: Plasma Concentrations of Steroid Hormones, Spontaneous and Induced Ovulation, and Quality of Eggs

The involvement of photoperiod and temperature in the regulation of reproductive processes was investigated in female yellow perch. Initially, all fish kept indoors were exposed to the same water temperature (22°C) and photoperiod (15L:9D). By the end of August, following the first sampling, fish were submitted to different photothermal regimes. Group A₃ was maintained under photothermal conditions characteristic for southern Ohio. Group B₃ was submitted to a condensed light/temperature regime designed to accelerate photothermal changes. The mean gonadosomatic index in group A₃ gradually increased throughout the experiment. Gonadosomatic index in group A₃ was higher than that in group B₃ from February through April. The highest plasma concentrations of estradiol in group A₃ occurred in November and December and exceeded those in group B₃. Plasma concentrations of testosterone in group A₃ were highest in December and tended to be higher in October and April than in other months. There was no difference in plasma testosterone between groups A₃ and B₃ except in March when concentrations of this androgen were extremely high in group B₃. Ovulation occurred earlier in B regime fish in comparison to that of A regime fish. However, egg quality of fish from regime B was lower than that of fish from regime A. We conclude that the condensed photothermal cycle is not an entirely effective method of inducing out-of-season spawning in female yellow perch. These data suggest that although the compression of the photothermal cycle slightly accelerated spawning, it also caused disturbances in patterns and levels of plasma steroids, diminished ovarian and hepatic growth, and reduced the quality of eggs. Thus, temperature and/or photoperiod may have only a modifying effect on the onset of perch spawning, but these environmental cues seem to markedly influence earlier stages of gonadal growth and development.     

Sirococcus shoot blight on Picea spinulosa in Bhutan

During a recent survey of forest tree diseases in Western and Central Bhutan, Sirococcus shoot blight and an associated Sirococcus sp. were found on saplings and mature trees of Eastern Himalayan spruce (Picea spinulosa). Based on morphological characteristics and DNA sequence comparisons of the ITS region of the rDNA operon, representative isolates from Bhutan were unequivocally identified as Sirococcus conigenus. The DNA sequence data also showed that these isolates belong to the P group of S. conigenus. To our best knowledge, this is the first report of Sirococcus shoot blight from the Himalayas or any other part of Asia. Sirococcus conigenus does not appear to cause dramatic damage at the moment, but this fungus has the potential to cause severe disease problems on P. spinulosa in Bhutan.     

Active metabolism in larval and juvenile fish: ontogenetic changes,effect of water temperature and fasting

Oxygen consumption, ammonia excretion and fish swimming speed were measured in fish induced to swim by optomotor reaction in a circular metabolism chamber. The relationship between the swimming speed and fish metabolism described by exponential equations allowed the extrapolation to the standard metabolism, i.e. at zero swimming speed. The partitioning of the catabolised protein in the energy supply was estimated based on AQ (volume of ammonia/ volume of oxygen) values. Weight specific standard metabolism, as expressed by the ammonia excretion rate, decreased by one order of magnitude in coregonids as the fish grew from 20 to 780 mg body weight. The slope of the relationship between oxygen uptake and swimming speed decreased in coregonid ontogenesis. In salmon, after 12 days of fasting 28% of energy used was derived from protein, whilst coregonid juveniles utilized mostly lipid. Active swimming in fasted juveniles of coregonid, as well as in salmon, led to the accelerated utilization of protein as a source of energy, based on AQ coefficients. In juveniles acclimated to a range of water temperatures from 14 to 26°C, the changes in standard or active metabolic rate (expressed as oxygen uptake or ammonia excretion) were described by Q₁₀ coefficients. They were generally higher for the ammonia excretion rate than for the oxygen uptake rate and for active metabolism than for standard metabolism. Utilization of protein as energy for swimming differed significantly between the species, being in general one order of magnitude higher in coregonids than in salmon. The use of protein for swimming activity tended to decrease during coregonid ontogenesis.     

Effects of ascorbic acid enrichment by immersion of rainbow trout (Oncorhynchus mykiss, Walbaum 1792) eggs and embryos

This study was conducted to examine the effects of different forms and concentrations of ascorbic acid (vitamin C), and different enrichment times (24 and 48 h post ovulation) on egg, embryo and alevin ascorbate concentrations and survival of rainbow trout (enrichment was at the ova stage). In experiments 1 and 2, fertilized eggs were immersed in water containing ascorbate at 0 (control), 100, 1000 mg L^?1 l ‐ascorbic acid (AA) and 2000 mg L^?1 l ‐ascorbyl monophosphate (AP). In experiment 3, 0 (control), 500 and 1000 mg L^?1 AA neutralized (N) with NaOH, 1000 mg L^?1 AA non‐neutralized (NN), 1000 and 2000 mg L^?1 AP immersions were used. The mean total ascorbic acid (TAA) and dehydroascorbic acid (DHA) concentrations were measured before fertilization, at 3 and 24 h after fertilization, at the eyed stage, and in hatched alevins. We observed significant differences in TAA concentration at different immersion levels at 3 and 24 h after fertilization. Survival decreased significantly depending on the level of vitamin C, pH of the solutions and immersion time. We suggest that when broodstock rainbow trout do not have enough vitamin C in their ovaries, immersion of eggs in 1000 mg L^?1 of neutralized AA may be useful.     

Protein requirements of grass carp fry (Ctenopharyngodon idella Val.)

The study was aimed at determining the optimal level of protein in a casein diet for grass carp. There was a linear relationship between the percentage of protein in the diet and the increase in fish protein and weight up to optimal levels of 41 and 43%, respectively. No further increase in fish protein or weight was noted with diets containing up to 64% of protein. However, if the optimal level was defined by the parabolic curve with the formula y = 36.21 + 6.238x ? 0.0593x², it would equal 52.6 ± 1.93%. Net protein utilization (NPU) values for experimental diets were obtained using a formula which took the maintenance nitrogen metabolism on a non-protein diet into consideration. This NPU formula was related to the length of the experiment, difference in the weight of fish at the beginning of the experiment, and growth of fish fed protein diets. Both protein efficiency ratio (PER) and NPU decreased with the increase in protein content of the diets according to the formulae y = 1.966?0.018x and $\text{y = 40.8?0.327x +}\text{302.6}\text{x}$, respectively.     

Effects of dietary cottonseed meal protein level on growth, gonad development and plasma sex steroid hormones of tropical fish tilapia Oreochromis sp

Five experimental diets containing increasing proportions of cottonseedmeal (CSM) protein (0, 25, 50, 75 and 100%; diets 1 to 5, respectively) toreplace fish meal (FM) protein were formulated for intensive culture of tilapiaOreochromis sp. Each diet was fed to three replicategroupsof fish (mean weight ± SE = 11.3 ± 3.9 g) in30L aquaria connected as a closed recirculating-water system andmaintained at 27 ± 1 °C. Fish were fed three times adayby hand at a rate of 3% of body weight during four weeks, after which thefeeding rate was gradually decreased to reach 1.5% at 16 weeks. Thesubstitutionof 75 and 100% of FM proteins by CSM proteins resulted in significantly lowerbody weights in both sexes. In both sexes, gonadosomatic indexes and plasmaconcentrations of sex steroids (testosterone, 11-ketotestoterone,estradiol-17 and 17,20-dihydroxy-4-pregnen-3-one) were notsignificantly different among dietary treatments. The concentration ofgossypol,an antifertility agent contained in CSM, was measured in reproductive tissues.The total gossypol concentration in the testis was consistently lower than thatmeasured in the ovaries of the same group. Moreover, in both sexes, theconcentration of the (+)isomer of gossypol was always higher than that of(–)isomer. The total gossypol concentrations in testes increasedsignificantly with the increase of CSM in the diet. The highest levels of the(+)isomer (7.64 ± 0.62 g g^–1)were found in the testes of fish fed diet 4, whereas the (–)isomerreached its highest values in the testes of fish fed diet 5. The highest levelsof both enantiomers of gossypol were found in the ovaries of fish fed diet 4(14.2 ± 2.7 and 5.6 ± 1.5 g g^–1for (+) and (–)isomers, respectively). In both sexes, thehistological analysis of the gonads did not reveal differences among the fishfed different levels of CSM. Although CSM at any levels did not affect thereproductive parameters examined in this study, it cannot be used to substitutemore than 50% of FM since at higher levels growth of tilapia was compromised.