Pronephric leucocytes of Cyprinus carpio: isolation, separation and characterization

Since the teleost pronephros is an important source of diverse immunocytes, suspensions of pronephric cells from young adult carp have been characterized. In freshly prepared suspensions, adherent, spreading cells (macrophages?) constituted less than 3% of the total population. Granulocytes and lymphocytes were co-dominant (less than 80%) leucocyte types. Continuous Percoll density gradient centrifugation yielded discrete subpopulations with these rho values and cytological characteristics: Fraction I & II rho = 1.055-1.070 thrombocytes, monocytes, macrophages, and lymphocytes. Fraction III rho = 1.080-1.090 granulocytes, type 1. Fraction IV rho = 1.105-1.110 erythrocytes and granulocytes, type 2. Fraction V rho = 1.118-1.125 granulocytes, type 3. Fraction VI rho = 1.140-1.150 granulocytes, type 4. Granulocyte motility increased markedly over the first 24 hr in vitro, and was enhanced by components washed from intact yeast. The subtypes of granulocytes were distinguishable by not only the rho values, but also on the basis of cell size, ultra-structure of the granules, and their histochemical and phagocytic characteristics. After simultaneous in vivo injection of Bacillus megaterium (Gram + ve), Aeromonas hydrophila (Gram - ve) and Saccharomyces cerevisiae (yeast), individual pronephric leucocytes were found capable of phagocytosing all three types of particle. Granulocytes which had phagocytosed B. megaterium were slower than macrophages in their ability to kill the bacteria. Encounter with B. megaterium or S. cerevisiae in vitro elicited a clumping reaction which involved mostly the larger leucocytes [granulocytes]. Both adherent cells and non-adherent cells were phagocytic in vitro.     

The effect of isolation and separation on the metabolism of sheep

For experimental purposes sheep are often isolated and/or separated from their conspecifics. In an experiment carried out with 12 Flevolander sheep which were put on a standard ration, we investigated how sheep react to the shift from a loose social housing system to a confined isolated environment (respiration chamber) where they remained for at least 10 days. Moreover we determined how many shifts it takes before the metabolism of the animal is completely adapted and no longer under stress in the less ideal environment.The main and most sensitive reaction after abrupt isolation is a significant increase in metabolic rate as estimated by heat production. Further, the digestive system reacts with a significant decrease in apparent digestibility. The haematocrit (PCV) increased significantly after isolation. All the analysed or calculated physiological parameters on the apparent digestibility of dry matter (DM), crude protein (CP) and energy (E) and on energy balance (EB), heat production (HP) and haematocrit (PCV) are significantly different for all sheep and periods at the 0.01 level. When the animals were moved for the 4th time into the respiration chamber, the apparent digestibility of the standard ration, the EB and PCV did not differ significantly from the consecutive periods. For metabolic rate (HP) we consider the sheep adapted in the 5th period.The economic importance of keeping sheep under stress free conditions is clearly demonstrated.     

Sheep peripheral blood-T-lymphocytes: isolation, separation and surface receptors for Helix pomatia agglutinin and peanut agglutinin from Arachis hypogaea

A study was made to develop and evaluate techniques for the isolation of ovine peripheral blood lymphocytes (PBL) and to investigate the suitability of peanut agglutinin (PNA) and Helix pomatia agglutinin (HP) as markers for sheep T-cells. The results show that sheen PBL can be prepared reproducibly by incubating ovine blood with carbonyl iron, centrifuging the blood over Percoll (colloidal silica polyvinylpyrrolidine) separating media (Pharmacia, Sweden) and harvesting the PBL at the interface. PBL so prepared, rarely undergo spontaneous agglutination, which is frequently seen with buffy coat cells and peripheral blood mononuclear cells. PNA and HP can be used as markers for ovine T-lymphocytes, because, under appropriate conditions, these lectins do not bind to B cells. Highly enriched peripheral blood T-lymphocytes were successfully prepared by the nylon wool technique and affinity column chromatography using HP. Highly purified B-cell sub-populations could not be prepared using the HP-Sepharose-6MB chromatography columns.     

蚕病病原菌种的衰退、复壮和保藏

根据常期从事蚕病病原微生物研究工作的经验,总结蚕病菌种在保藏中常出现的衰退现象,介绍防止衰退的主要措施。     

Studies on the pathogenicity ofBabesia bovis in water buffaloes after cryopreservation and resuscitation

Tropical Animal Health and Production - Packed erythrocytes infected withBabesia bovis were mixed with an equal volume of 16% dimethyl sulphoxide (DMS0) in Alsever’s solution and dispensed...     

The isolation, propagation and characterization of tissue-cultured equine rotaviruses

From 105 field cases of diarrhea in neonatal or young foals, rotavirus was detected by electron microscopy (EM) and/or by enzyme-linked immunosorbent assay (ELISA) in the feces of 65 foals on 16 different premises. ELISA was performed with Rotazyme test kits developed by Abbot and Company for the detection of rotaviruses. Twenty-four field isolates from the feces of diarrheic foals with equine rotavirus infection as ascertained by EM were placed in MA-104 cell cultures after pretreatment of the viral suspension with 10 micrograms ml-1 of trypsin and incorporation of 0.5 micrograms ml-1 or 1 microgram ml-1 of trypsin in Earle's minimal essential medium (MEM), 2% lactalbumen hydrolysate, and antibiotics. The isolates that replicated in cell culture produced varying degrees of cytopathic effect. After the 24 isolates had been transferred 5 or 7 times in cell culture, viral particles were observed in 17 by EM, and 22 had positive ELISA tests as determined by visual color chart and spectrophotometric readings. Concentrated tissue-cultured viral antigen of 9 isolates fixed complement using Nebraska calf diarrhea rotavirus calf antiserum while four isolates gave negative results. The same 13 tissue-cultured viral suspensions failed to fix complement using reovirus antiserum. The 9th passages of two isolates (EID1 and EID2) yielded titers of 10(4.45) ml-1 TCID50 and of 10(4.95) ml-1 TCID50, respectively, as measured by cytopathic effect. After 13 tissue-cultured passages, 2 other isolates, EID3 and EID4, each had titers of 10(6.2) ml-1 TCID50 and of 10(5.95) ml-1 TCID, respectively. Cytoplasmic or intranuclear inclusions were not seen in any cells of the MA-104 infected cell cultures. Small, but distinct, plaques in MA-104 cell cultures were produced by the EID1 isolate. Polyacrylamide gel electrophoresis tests of EID1 and EID2 isolates at the 9th cell passage and EID3 and EID4 isolates at the 13th cell passage each showed that the RNA genome had 11 segments with a migrating pattern that was identical for each isolate and characteristic of rotaviruses. These 4 equine tissue-cultured isolates when tested by ELISA, utilizing a monoclonal antibody serum pool that cross-reacted with many rotavirus isolates, each gave positive values comparable to rotavirus antigen controls.     

The immune responses to antigenic variants of Babesia bigemina in the bovine.

Four Babesia bigemina stabilates were used to determine the immune response of cattle to acute and chronic blood- and tick-borne infections. Thirty-two intact calves were divided into 16 groups of two and each calf was inoculated with infective B bigemina erythrocytic stabilates. Twenty-eight days later they were challanged with homologous and heterologous stabilates, and monitored for an additional 20 days. The hosts' apparently reduced response to homologous challenge but marked immune response to heterologous challenge indicated antigenic differences between the isolates and confirmed the conclusions reached by examination of the serological data.     

波尔山羊耳成纤维细胞的分离培养与保存

本文研究了波尔山羊耳成纤维细胞的体外培养,包括分离纯化和生长特征,同时还研究了成纤维细胞的冷藏和冷冻保存.结果表明,波尔山羊耳成纤维细胞贴壁生长,一般经历潜伏期、指数增生期和平台期;4℃体外冷藏时间为5 d;冷冻保存时4℃平衡1 h、-80℃过夜,然后直接投入液氮中,解冻后培养得到61.6%的贴壁率.     

The isolation, characterisation and quantification of the equine plasma lipoproteins

Plasma lipoproteins were isolated from eight Thoroughbred horses and eight Shetland ponies on the basis of particle size by gel filtration chromatography and according to density using rate-zonal ultracentrifugation. Three major classes corresponding to very low density lipoproteins (VLDL), low density lipoproteins (LDL) and high density lipoproteins (HDL) were identified and characterised by their lipid and apolipoprotein compositions. The particle size distributions of each class were determined by electron microscopy and non-denaturing polyacrylamide gradient gel electrophoresis. HDL was found to dominate the equine lipoprotein spectrum, accounting for 61 per cent of the total plasma lipoprotein mass (VLDL 24 per cent, LDL 15 per cent). The VLDL class was isolated as a single population of particles that were triglyceride rich and cholesterol, phospholipid and protein poor. Equine LDL was characteristically cholesterol rich and was found to be polydisperse comprising three subfractions that were discrete with respect to particle size and lipid composition. The HDL class was composed of homogeneous particles that were typically protein rich. Apolipoprotein (apo) B was the major protein of VLDL and LDL, and presented two components on polyacrylamide gel electrophoresis with molecular weights in the region of human apoB-100 and a third in VLDL similar to that of apoB-48. ApoA-I was the predominant protein in equine HDL. Although there were no breed differences in the physical or chemical properties of each lipoprotein class, the Shetland ponies had higher plasma triglyceride and VLDL concentrations than their Thoroughbred counterparts.     

The effect of neutrophils, tumor necrosis factor, and granulocyte macrophage/colony stimulating factor on Babesia bovis and Babesia bigemina in culture.

Bovine neutrophils, human recombinant tumor necrosis factor-alpha (TNF), and bovine recombinant granulocyte macrophage/colony stimulating factor (GM/CSF) were added to microaerophilic cultures of Babesia bovis and Babesia bigemina to determine if those substances could inhibit growth. Incorporation of [3H]hypoxanthine by the Babesia spp. was utilized as an indirect measure of parasite growth. When neutrophils were added to cultures of B. bovis and B. bigemina, the highest percentage inhibition of growth was attained. There was no significant enhancement of neutrophil killing when TNF or GM/CSF or both were added to either Babesia spp. Addition of TNF or GM/CSF or both substances (without neutrophils) resulted in an increase in growth of B. bovis and B. bigemina. For B. bovis, the group that contained neutrophils only and the group that contained neutrophils and TNF resulted in significantly higher growth inhibitions than the treatment group which contained neutrophils and GM/CSF or the group that contained neutrophils, TNF, and GM/CSF. No significant differences in inhibition were observed for the same treatment groups between B. bovis and B. bigemina.     

Prevalence of Anaplasma marginale, Babesia bigemina and B. bovis in Nigerian cattle using serological methods

Serum samples collected randomly from 500 cattle from the 10 northern states of Nigeria were tested for antibodies against Anaplasma marginale by indirect fluorescent antibody (IFA), card agglutination (CT) and capillary tube-agglutination (CA) tests. The serum samples were also examined for antibodies to Babesia bigemina and B. bovis by the IFA test only. Of the serum samples tested, 79.4% had antibodies against A. marginale by the IFA test, 40 and 25% in the CT and CA tests, respectively. The IFA test results for B. bigemina and B. bovis were 29.4 and 14.1%, respectively.