Studies of Enteric Pathogens and γ-Globulin Levels of Neonatal Calves in Sweden

Faecal and blood samples were taken from 10-30% of calves, 36 hours to 14 days old, in 47 dairy herds in different regions of Sweden from September 1987 to October 1988 (Olsson et al 1993). Faecal samples from 279 calves were analysed for the presence of Escherichia coli (K99+), rotavirus and Cryptosporidium sp. Twenty (7.2%) of these samples were from diarrhoeic calves. An ELISA was developed and used for the rotavirus analysis. E. coli K99+ was detected in 11.5%, Cryptosporidium sp. in 6.1% and rotavirus in 5.4% of the faecal samples. The presence of rotavirus alone and the combination rotavirus and E. coli (K99+) was found to be associated with diarrhoea (p<0.001 and p<0.01 respectively). Blood samples from 327 calves were analysed for the level of total protein and γ-globulin. In 43 of these samples (13%) γ-globulin did not separate from the ß2-region by electrophoresis. The mean total protein concentration was 53.6 g/1 in calves free from diarrhoea. The mean γ-globulin concentration, adjusted to 7 days age was 5.9 g/1. The 20 diarrhoeic calves had lower levels of both total protein and γ-globulin, compared with calves without diarrhoea, but the difference was not significant. One litre more of colostrum at the first feed increased the level of total protein of the calves’ sera by 1.4 g/1 (p = 0.05). Calves born between May and September had a 2.0 g/1 higher (p<0.001) serum concentration of γ-globulin than calves born between October and April.     

Plasma PGF2α metabolite levels in cats with uterine disease

Uterine disease induces PGF_2α increase in many animal species, which can be measured by the metabolite 15-keto-(13,14)-dihydro-PGF_2α (PGFM). Plasma PGFM levels are associated with severity of the uterine disease and presence of systemic inflammatory response syndrome (SIRS) in dogs. The objectives in this study were to investigate PGFM levels, presence of SIRS, and clinical and laboratory parameters in female cats as possible indicators for severity of uterine disease. In total, 7 female cats with pyometra, 2 with mucometra, 7 with cystic endometrial hyperplasia (CEH), and 14 healthy control cats were included. Physical examination, ovariohysterectomy, and histopathology were performed, laboratory parameters were analyzed, and PGFM levels were analyzed by radioimmunoassay. Analysis of variance, Fisher's exact test, Student's t-test and Pearson's product moment correlation coefficient were used for data analysis. In cats with pyometra, mean PGFM levels were increased (21.1 nmol L⁻¹) but were decreased in cats with CEH (0.4 nmol L⁻¹) compared with control cats (0.6 nmol L⁻¹). In cats with mucometra, the mean PGFM level was 8.8 nmol L⁻¹. Systemic inflammatory response syndrome was present in 6 (85%) cats with pyometra, 1 cat with mucometra, and 1 cat with CEH. Hospitalization length was negatively correlated with albumin and positively correlated with total white blood cell count (WBC), neutrophils, band neutrophils (BN), percentage BN (PBN), and monocytes. Pyometra and mucometra were associated with increased plasma levels of PGFM. The parameters albumin, WBC, neutrophils, BN, PBN, and monocytes may be useful to determine morbidity as measured by hospitalization length.     

Immunochemical Isolation of γ-Globulin mRNA and Estimation of Immunoglobulin Gene Reiteration

The polyribosomes synthesizing γ-globulin have been isolated by the achievement of specific precipitation using bentonite-treated anti-IgG antibody. The RNA extracted from the immunochemically precipitated polysomes was tested for its ability to direct the synthesis of proteins in a cell-free system. The specific γ-globulin-synthesizing activity (cpm of γ-globulin synthesized/μg RNA) of this RNA was 10-fold greater than that from total polysomes. γ-globulin mRNA (messenger RNA) isolated by immunoprecipitation was more than 89% pure with respect to contamination by other species of mRNA. The products synthesized by the cell-free system were also analyzed by sodium dodecyl sulphate(SDS)-polyacrylamide gel electrophoresis. This RNA has been hybridized with mouse myeloma DNA. The estimation of immunoglobulin gene reiteration was carried out using hybridization kinetics with consideration given to the DNA/RNA ratio since the estimation from the “half Cot value” is not accurate. The results suggest that in the mouse there are about 20 copies per subgroup of genes coding for the variable region of the H and L chains.     

An extracellular 5'-nucleotidase with both monoesterase and diesterase activity from Micrococcus sodonensis.

Bovine γ-globulin was separated into four fractions by chromatography on cellulose phosphate. The chromatographic distribution was similar to that reported for human and dog γ-globulin. More than 80% of a nonspecific phagocytosis stimulating factor (leucokinin) present in the serum was isolated in γ-globulin fraction IV. Bocine red blood cells and polymorphonuclear leukocytes bind γ-globulin without appreciable selectivity for any of the four chromatographic fractions, but they do selectively bind the phagocytosis stimulating factor. Splenectomy caused no observable change in either the chromatographic distribution or phagocytosis stimulating activity of bovine serum γ-globulin. The tetrapeptide tuftsin stimulates phagocytosis by bovine neutrophiles, but on a molar basis the activity of tuftsin was only 10% that of the phagocytosis stimulating factor. If the factor exerts its effect, as has been proposed, by having a phagocytosis stimulating peptide cleaved from it by an enzyme on the leukocyte membrane, that peptide must differ in structure from tuftsin. This conclusion is supported by the inability of trypsin to liberate an active peptide from bovine serum.     

Hyperferritinemia and Hyperuricemia May Be Associated with Liver Function Abnormality in Obese Adolescents

Background

The iron status in human body and its association with liver function in adolescents was rarely studied. The objective was to investigate the association among the levels of serum ferritin, uric acid and alanine aminotransferase (ALT) in adolescents.

Methods and Results

A total of 2090 adolescents negative for hepatitis B surface antigen from one junior high school (786, 12–13 years), three senior high schools (973, 15–16 years) and one college (331, 18–19 years) participated in this survey. Anthropometric and biochemical measurements, including complete blood count, ALT, serum ferritin and uric acid were performed. An ALT>42 U/L was defined as elevated, a ferritin level >200 µg/L was defined as hyperferritinemia. A uric acid level >460 µmol/L in males and >340 µmol/L in females was defined as hyperuricemia. The chi-squared test, linear regression and multivariate logistic regression were used for the data analysis. Elevated ALT levels were detected in 76 (3.6%) students and were more prevalent in males than females (6.4% vs. 2.0%, p<0.001). The univariate analysis found gender, age group, body mass index, ferritin level, uric acid level and white blood cell count all to be significantly associated with elevated ALT. Linear regression showed a positive correlation among log(ferritin), uric acid level and ALT level. Elevated ALT occurred more frequently at ferritin level >100 µg/L. The logistic regression analysis found that body mass index, hyperferritinemia and hyperuricemia were significant factors associated with the ALT elevation, but gender, age, and white blood cell count were not.

Conclusions

Hyperferritinemia and hyperuricemia are two independently significant factors associated with ALT elevation among obese adolescents. More studies are needed to corroborate any hypothesis related to these phenomena.     

Genetic and environmental causes of variation in basal levels of blood cells.

The genetic and environmental determinants of variation in blood cell size and number were investigated in 392 pairs of 12-year-old twins. The following blood cell indices were measured: haemoglobin, red blood cell count, haematocrit, mean corpuscular volume, platelet number, total white cell count, level of neutrophils, monocytes, eosinophils, total lymphocytes, CD3+ lymphocytes, CD4+ lymphocytes, CD8+ lymphocytes, CD19+ lymphocytes, CD56+ lymphocytes and CD4+/CD8+ ratio. Genetic factors contributed significantly to all blood cell measures accounting for between 61 and 96% of variance. Heritability estimates did not differ significantly between males and females, although the sample size of the present study was not large enough to exclude the possibility of sex-limited gene expression. Common environmental factors were important in determining red blood cell count and haematocrit, but were not important in determining basal levels of any white blood cell type.     

Agarose Gel Electrophoretic Fractionation of Serum Proteins in Adult Cattle. II. A Study Of Cows with Different Diseases

In 283 dairy cows suffering from internal disorders the serum proteins were studied by agarose gel electrophoresis supplemented by total protein and albumin determinations. The clinical diagnoses could be grouped according to protein pattern. Group 1 (abomasal displacement and traumatic muscle injury) did not appreciably affect the serum protein concentrations and represented primarily non-inflammatory diseases or diseases of non-infectious origin. Group 2 (leukosis) occupied an exceptional position, with heavy lowering of albumin unaccompanied by a corresponding rise of total globulin. The γ-globulin concentration was significantly lowered. Group 3 (acute traumatic peritonitis) represented an acute inflammatory process with increase chiefly of the α-globulins, while the γ-globulin concentration was normal. Group 4 (chronic traumatic peritonitis, summer mastitis, chronic subclinical mastitis, chronic laminitis and polyarthritis, urinary tract infection, abscess, and sub-acute-chronic pneumonia) comprised diseases chiefly of chronic inflammatory character and with infectious origin. Especially characteristic was the heavy rise of globulins in general and of γ-globulin in particular. In most cases there was a large increase also of the α- and p-globulin fractions.     

White blood cell count and adherence in sportsmen and non-training subjects

The white blood cell count and differential white blood cell count as well as the adherence of these cells were determined in 31 young clinically healthy men not participating actively in any sports and in 32 footballers belonging to third league sports clubs. In the group of footballers the determinations were carried out twice: in the initial period and then at the end of the highest-form period of the one-year training cycle. In the footballers in relation to the control group the total white blood cell count was raised, the differential count showed a shift in favour of neutrophils, eosinophils and monocytes at the expense of lymphocytes. The lymphocyte adherence was significantly decreased.     

Aerobic Bacteria Occurring in the Vagina of Bitches with Reproductive Disorders

A retrospective survey was performed of aerobic bacterial species found in the vagina of 203 bitches with genital disorders, e.g. infertility, vaginitis, pyometra and puppy death. Escherichia coli, beta-hemolytic streptococci, Staphylococcus intermedius and Pasteurella multocida were the species most often isolated. From bitches with pyometra E. coli in pure culture was the most frequent isolate. In contrast, the majority of infertile bitches gave rise to mixed cultures, and no specific bacterial species was consistently associated with infertility. Thus, bacterial sampling from infertile bitches was concluded to be of low diagnostic value. Bacterial species isolated from the bitches having vaginitis were present in pure culture in 26.9% of the samples while nonspecific mixed cultures were obtained from 34.6% of the samples from these bitches. E. coli was the most frequently isolated bacterial species from bitches with dead puppies. However, in such cases it is important to relate the vaginal bacterial findings to autopsy findings and the results of bacteriological cultures of the pups.     

Different bacterial lipopolysaccharides as toxicants and stressors in the shrimp Palaemon elegans

This study compares the in vivo haemocytic response of shrimp, Palaemon elegans (Rathke) to different types of LPS injection. In particular it investigates to what degree and speed the haemocytopenia varies between LPSs from different sources. It further compares the tolerated doses of different LPSs in these animals and finds substantial differences in the various toxicity types. The work then relates this to blood glucose levels and stress-linked variations in glycaemic status. The order of LPS decreasing toxicity determined by LD50 at 96 h was: Salmonella enteritidis, Serratia marcescens, Pseudomonas aeruginosa 10, Escherichia coli K-235 and E. coli 0111:B4. Eyestalkless animals were more sensitive to LPS. The effects of injected LPS on circulating total blood cell count (THC) was tested. The results show that LPS caused a decrease in THC 8 h after injection and then the THC returned to the initial level and this effect depended on the LPS tested. E. coli K-235 was the most effective in causing haemocytopenia followed by E. coli 0111:B4, S. enteritidis, S. marcescens, and P. aeruginosa 10. Moreover, LPS-induced increases in the blood glucose level and the time and dose related curves of response obtained depended on the type of LPS tested. E. coli K-235 LPS was again the most effective in elevating blood glucose followed by E. coli 0111:B4, S. marcescens, S. enteritidis and then P. aeruginosa 10. No significant hyperglycaemia was induced in eyestalkless animals. An inverse order relationship between toxicity (LD50) and stress responses (hyperglycaemia and THC decrease) may suggest a defensive and adaptive role of the latter in occasional septicaemia.     

National field evaluation of a defined substrate method for the simultaneous detection of total coliforms and Escherichia coli from drinking water: comparison with presence-absence techniques.

A defined substrate method was applied to drinking water to simultaneously enumerate total coliforms and total Escherichia coli directly from samples. After incubation at 35 degrees C for 24 h, the development of yellow in an initially colorless solution was specific for total coliforms; fluorescence at 366 nm in the same tube(s) or vessel demonstrated the presence of E. coli. No confirmatory or completed steps were necessary. Known as autoanalysis colilert (AC), this method was constituted as a presence-absence test and compared with the methods described in Standard Methods (SM) in the P-A format. Seven water utilities representing a wide geological and hydrological spectrum participated in the evaluation. A total of 702 split drinking water samples were analyzed. Of these, 358 were negative in both tests (SM- and AC-); 302 were positive (SM+ and AC+); and 42 were mixed (SM+ and AC-, 20; AC+ and SM-, 22). The overall agreement rate was 94%. Comparison of the SM and AC results by nonparametric statistics demonstrated no differences. Heterotrophic plate count bacteria exerted no discernible effect on the AC test. By subculture, each time the AC test was yellow, a total coliform was present; when the test was fluorescent, E. coli was isolated.     

National field evaluation of a defined substrate method for the simultaneous detection of total coliforms and Escherichia coli from drinking water: comparison with presence-absence techniques

A defined substrate method was applied to drinking water to simultaneously enumerate total coliforms and total Escherichia coli directly from samples. After incubation at 35 degrees C for 24 h, the development of yellow in an initially colorless solution was specific for total coliforms; fluorescence at 366 nm in the same tube(s) or vessel demonstrated the presence of E. coli. No confirmatory or completed steps were necessary. Known as autoanalysis colilert (AC), this method was constituted as a presence-absence test and compared with the methods described in Standard Methods (SM) in the P-A format. Seven water utilities representing a wide geological and hydrological spectrum participated in the evaluation. A total of 702 split drinking water samples were analyzed. Of these, 358 were negative in both tests (SM- and AC-); 302 were positive (SM+ and AC+); and 42 were mixed (SM+ and AC-, 20; AC+ and SM-, 22). The overall agreement rate was 94%. Comparison of the SM and AC results by nonparametric statistics demonstrated no differences. Heterotrophic plate count bacteria exerted no discernible effect on the AC test. By subculture, each time the AC test was yellow, a total coliform was present; when the test was fluorescent, E. coli was isolated.     

Grafting of different glycosides on the surface of liposomes and its effect on the tissue distribution of 125I-labelled γ-globulin encapsulated in liposomes

Different glycosides were grafted on the surface of liposomes containing ¹²⁵I-labelled γ-globulin by two ways: (1) by using glycolipid and (2) by covalent coupling of $\text{p-}\text{aminophenyl-}\text{d}\text{-glycosides}$ to phosphatidylethanolamine liposomes using glutaraldehyde. The distribution of ¹²⁵I-labelled γ-globulin was determined in mouse tissues from 5–60 min after a single injection of these liposomes. The liver uptake of encapsulated ¹²⁵I-labelled γ-globulin was highest from liposomes having galactose and mannose on the surface. Competition experiments and cross-inhibition studies indicate that this uptake are mediated by specific recognition of the surface galactose and mannose residues of liposomes by the receptors present on the plasma membrane of liver cells. Stearylamine-containing liposomes were found to be more efficient in mediating the uptake of ¹²⁵I-labelled γ-globulin by the lung, whereas in the case of spleen, phosphatidylethanolamine liposomes were more efficient. The extent of uptake of ¹²⁵I-labelled γ-globulin from all types of liposome decreases as the amount of given liposomes increases. The uptake of ¹²⁵I-labelled γ-globulin from liposomes containing asialogangliosides depends upon the phospholipid/ glycolipid ratio. These experiments clearly demonstrate that enhanced liposome uptake by liver cells could be achieved by grafting galactose and mannose on the liposomal surface.     

Hematologic values of New Zealand white rabbits determined by automated flow cytometry.

Hematologic values of peripheral blood from normal adult New Zealand White rabbits were determined by five different automated flow cytometers in use in a routine clinical hematology laboratory: Technicon H1, Coulter Counter CC540, Coulter Counter VCS, Sysmex NE8000 and Sysmex R1000. The software designed for human blood analysis was used in all instances without adaptation. The total numbers of white blood cells, red blood cells, reticulocytes and platelets were measured with high precision and accuracy. Except for hemoglobin content, concordance was excellent for all measured and calculated values among the different automated flow cytometers. Determining the white blood cell differential count was more complex. Eosinophils and lymphocytes were quantified reliably by all the automated flow cytometers used. However, the results were rejected by Technicon H1 and Sysmex NE8000 in 50% of the cases. Rabbit basophils were recognized with accuracy by Technicon H1 only. The proportion of polymorphonuclear versus mononuclear white cells was identical when measured with Technicon H1 and Coulter Counter VCS. These results show that the new generation of automated flow cytometers designed for human blood can be used with some limitations for animal studies. They allow the standardization of normal values and comparison of results among or between laboratories. They also introduce new parameters, the value of which is as yet undefined.     

Clinico-laboratory substantiation of possible use of the indirect hemagglutination test for the interpretation of diagnosis of enteropathogenic E. coli carrier state]

The diagnostic value of the indirect hemagglutination (IHA) test in deciphering the diagnosis of enteropathogenic E. coli carrier state was found in this work; this was done by comparing the specific antibody level in carriers and patients suffering from coli-infection and also by establishing the correlation between the results of the IHA test and the clinical and laboratory examination of carriers.     

Influences of the interference of γ-tubulin gene expression on the morphology and microtubules of ciliate Euplotes eurystomus

A recombinant plasmid L4440-GTU expressing γ-tubulin dsRNA of the ciliate Euplotes eurystomus was constructed and transferred into Escherichia coli strain HT115. The resultant E. coli bacteria were fed on E. eurystomus to inhibit the ciliate's γ-tubulin gene expression. As a result, the γ-tubulin gene expression level was decreased, and this inactivation blocked cell division, which was lethal. In addition, the loss of the C-tubule from the nine-microtubule triplets in basal bodies, and the disappearance of some microtubules or mislocalization of some microtubule organization units in the subpellicular microtubule layers were also observed. These results indicate that the γ-tubulin is not only important for the stability of the nine-microtubule triplets in basal bodies, but also necessary for the integrity of microtubule organization patterns in the subpellicular microtubule layer.     

人白细胞介素18的基因克隆与表达

人白细胞介素１８（ＩＬ－１８）是新近发现的细胞因子之一．研究表明它参与Ｔ１辅助细胞介导的细胞免疫．利用ＲＴ－ＰＣＲ技术从人外周血细胞扩增得到了ＩＬ－１８的ｃＤＮＡ并测定其核酸序列．利用基因重组技术构建ＩＬ－１８的表达载体,并在大肠杆菌中进行了表达．这为以后进一步研究ＩＬ－１８的功能奠定了基础．     

Comparison of Vero cell assay, polymerase chain reaction and an enzyme immunoassay for identification of verocytotoxin-producing Escherichia coli O157:H7

This study evaluated three different analytical methods for identification of Verocytotoxin-producing E. coli O157:H7 (VTEC) strains. A total of 34 E. coli O157:H7 strains isolated from bovine faeces and bovine carcasses were comparatively tested with Vero cell assay (VCA), PCR and the sandwich ELISA "RIDASCREEN Verotoxin" test. The VCA, performed without a neutralization assay, gave a false positive result because a VCA-positive E. coli O157:H7 strain did not possess the VT-coding genes when tested with PCR. The lack of specificity of the VCA could be avoided by testing for neutralization of cytotoxicity. The commercial ELISA system was as sensitive and specific as PCR, with the advantages of being a more rapid and easier procedure which could be employed in all first level diagnostic laboratories.     

Accuracy and Feasibility of Point-Of-Care White Blood Cell Count and C-Reactive Protein Measurements at the Pediatric Emergency Department

Background

Several point-of-care (POC) tests are available for evaluation of febrile patients, but the data about their performance in acute care setting is sparse. We investigated the analytical accuracy and feasibility of POC tests for white blood cell (WBC) count and C-reactive protein (CRP) at the pediatric emergency department (ED).

Methods

In the first part of the study, HemoCue WBC and Afinion AS100 CRP POC analyzers were compared with laboratory’s routine WBC (Sysmex XE-2100) and CRP (Modular P) analyzers in the hospital central laboratory in 77 and 48 clinical blood samples, respectively. The POC tests were then adopted in use at the pediatric ED. In the second part of the study, we compared WBC and CRP levels measured by POC and routine methods during 171 ED patient visits by 168 febrile children and adolescents. Attending physicians performed POC tests in capillary fingerprick samples.

Results

In parallel measurements in the laboratory both WBC and CRP POC analyzers showed good agreement with the reference methods. In febrile children at the emergency department (median age 2.4 years), physician performed POC determinations in capillary blood gave comparable results with those in venous blood analyzed in the laboratory. The mean difference between POC and reference test result was 1.1 E9/L (95% limits of agreement from -6.5 to 8.8 E9/L) for WBC and -1.2 mg/L (95% limits of agreement from -29.6 to 27.2 mg/L) for CRP.

Conclusions

POC tests are feasible and relatively accurate methods to assess CRP level and WBC count among febrile children at the ED.     

Seasonal hematology and serum chemistry of wild beluga whales (Delphinapterus leucas) in Bristol Bay, Alaska, USA

We collected blood from 18 beluga whales (Delphinapterus leucas), live-captured in Bristol Bay, Alaska, USA, in May and September 2008, to establish baseline hematologic and serum chemistry values and to determine whether there were significant differences in hematologic values by sex, season, size/age, or time during the capture period. Whole blood was collected within an average of 19 min (range=11-30 min) after the net was set for capture, and for eight animals, blood collection was repeated in a later season after between 80-100 min; all blood was processed within 12 hr. Mean hematocrit, chloride, creatinine, total protein, albumin, and alkaline phosphatase were significantly lower in May than they were in September, whereas mean corpuscular hemoglobin concentration, monocytes, phosphorous, magnesium, blood urea nitrogen, alanine aminotransferase, aspartate aminotransferase, γ-glutamyltranspeptidase, and creatinine kinase were significantly higher. Mean total protein, white blood cell count, neutrophils, and lymphocytes were significantly higher early in the capture period than they were later. No significant differences in blood analyte values were noted between males and females. Using overall body length as a proxy for age, larger (older) belugas had lower white blood cell, lymphocyte, and eosinophil counts as well as lower sodium, potassium, and calcium levels but higher creatinine levels than smaller belugas. These data provide values for hematology and serum chemistry for comparisons with other wild belugas.