Anaplasmosis in Uganda. I. Use of dried blood on filter papers and serum samples for serodiagnosis of anaplasmosis—A comparative study

Summary The suitability of blood collected on filter papers in comparison with corresponding conventional serum samples in the diagnosis of bovine anaplasmosis was studied using the complement fixation test, DOT-ELISA, Western immunoblot and rapid card agglutination test. Dried blood on Whatman filter paper no. 1 was eluted in PBS 0·05% Tween 20 giving an initial dilution of 1∶10. The reactivity of the eluted samples in both DOT-ELISA and Western immunoblotting were similar to those obtained with the corresponding straight serum sample dilutions. Filter paper samples gave lower reactivity in the remaining tests when compared with corresponding serum samples. There was no significant difference in the reactivity between the eluates from filter papers stored at temperatures ranging between 15·5 and 24°C and those kept refrigerated. Storage at 15·5 to 24°C did not significantly affect reactivity for up to six months. Eluates from filter papers stored for six months at 15·5 to 24°C continued to give similar reactivity as those from freshly prepared filter papers in both DOT-ELISA and Western blot, and in the rapid card agglutination test. It is concluded that collecting blood on filter papers is a suitable technique for large scale seroepidemiological studies on anaplasmosis and offers many advantages in developing countries where transport and cold chain facilities are a major constraint.

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Resumen Se estudió la efectividad de muestras de sangre colectadas en papel filtro, en comparación con las correspondientes muestras convencionales du suero, en el diagnóstico de anaplasmosis bovina, utilizando fijación de complemento, DOT-ELISA, Western, immunoblot y la prueba rápida de la tarjeta. La sangre seca en papel Whatman N^o 1 fue removida con PBS 0·05% entre 20, dando una dilución inicial de 1∶10. La reactividad de las muestras removidas de papel filtro, en la prueba de DOT-ELISA y Western immunoblotting, fueron similares a la obtenida con la correspondiente muestra de suero. Las muestras de papel filtro reaccionaron menos en las otras pruebas, cuando se compararon con las correspondients muestras du suero. No hubo diferencia significativa en la reactividad entre los lavados del papel filtro guardados a temperaturas entre 15·5 y 24°C y aquellos guardados en refrigeración. El almacenaje entre 15·5 y 24°C, no afectó la reactividad hastas seis meses. Los lavados de papel filtro guardados por seis meses entre 15·5 y 24°C, dieron la misma reactividad como los lavados frescos, en la prueba DOT-ELISA y Western blot, lo mismo que en la prueba de aglutinación rápida de tarjeta. Se concluye, qu la colección de sangre en papel filtro, es una buena técnica para estudios epidemiológicos de cierta magnitud, sobre anaplasmosis, ofreciendo ventajas considerables en paises en desarrollo en donde las cadenas de frío son deficientes.

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Résumé La fiabilité du sang récolté sur papier filtre comparée à celle des prélèvements conventionnels de sérum pour le diagnostic de l'anaplasmose a été étudiée à l'aide des tests suivants: fixation du complément, ELISA, immunoblot de Western, test rapide d'agglutination sur carte. Du sang séché sur papier filtre Whatman N^o 1 a fait l'object d'une élution dans une solution de PBS à 0,05 p. 100 (Tween 20) pour donner une dilution de base au dilution de base au 1∶10. Le réactivité des échantillons, autant avec le test ELISA que l'immunoblot Western, a été identique à celle obtenue par dilution directe de sérums homologue. Les échantillons sur papier filtre ont donné une réactivité plus faible pour les autres tests, comparée à celle des échantillons de sérum semblables. Aucune différence significative n'a été décelée quant à leur réactivité les éluats provenant de papiers filtres stockés à des températures comprises entre 15,5 et 24°C at ceux conservés au réfrigérateur. Le stockage entre 15,5 et 24°C n'a pas non plus affecté la réactivité de fa?on significative; les éluats conservés à partir des papiers filtres, à cette même température durant 6 mois, ont montré des réactions identiques que ceux provenant de papiers filtres fra?chement préparés, à la fois avec le test ELISA, celui de Western Blot et le test d'agglutination rapide sur carte. On peut conclure que la collecte du sang sur papier filtre est une technique adaptée à l'étude épidémiologique de l'anaplasmose à grande échelle. Elle offre de nombreux avantages dans les pays en développement où offre de nombreux avantages dans les pays en développement où les moyens de transports et la cha?ne du froid constituent des contraintes majeures.

     

Flow Cytometry Identification of X- and Y-Chromosome-Bearing Goat Spermatozoa

Flow cytometric sorting technology was used to measure the difference in DNA content between X- and Y-chromosome-bearing spermatozoa in bucks. Spermatozoa were analysed by flow cytometry to characterize X- and Y-chromosome-bearing sperm populations and to quantify the DNA difference between them. Two symmetrical, overlapping and clearly separated peaks, corresponding to X- and Y-bearing spermatozoa, were detected. The difference in fluorescence intensity between the peaks was 4.4 +/- 0.03% without any significant inter- or intra-animal variations. Therefore, the identification and selection of high-purity samples of sperm populations for sex sorting is easier in bucks compared with other domestic species.     

From regionally predictable to locally complex population structure in a freshwater top predator: river systems are not always the unit of connectivity in Northern Pike Esox lucius

Contemporary genetic diversity is the product of both historical and contemporary forces, such as climatic and geological processes affecting range distribution and continuously moulded by evolutionary forces selection, gene flow and genetic drift. Predatory freshwater fishes, such as Northern Pike Esox lucius, commonly exhibit small population sizes, and several local populations are considered endangered. Pike inhabit diverse habitat types, including lakes, rivers and brackish marine waters, thus spanning from small isolated patches to large open marine systems. However, pike population structure from local to regional scales is relatively poorly described, in spite of its significance to developing conservation measures. We analysed microsatellite variation in a total of 1185 North European pike from 46 samples collected across both local and regional scales, as well as over time, to address two overarching questions: Is pike population structure associated with local and/or regional connectivity patterns, and which factors likely have the main influence on the contemporary distribution of genetic diversity? To answer this, we combined estimators of population diversity and structure to assess evidence of whether populations within (i) habitats, (ii) drainage systems and (iii) geographical regions are closer related than among these ranges, and whether patterns are temporally stable. Contrasting previous predictions that genetic drift obscures signals of postglacial colonisation history, we identified clear regional differences in population genetic signatures, suggesting a major effect of drainage divides on colonisation history and connectivity. However, several populations deviated from the general pattern, showing that local processes may be complex and need to be examined case‐by‐case.     

Relative bioavailability of organic bis-glycinate bound copper relative to inorganic copper sulfate in beef steers fed a high antagonist growing diet

To assess the relative bioavailability of bis-glycinate bound Cu, 90 Angus-cross steers (265 ± 21 kg) were blocked by body weight (BW) to pens with GrowSafe bunks and randomly assigned to dietary treatments (14 to 18 steers/treatment): 0 mg supplemental Cu/kg dry matter (DM; CON), 5 or 10 mg supplemental Cu/kg DM as Cu sulfate (CS5; CS10) or bis-glycinate bound Cu (GLY5; GLY10). Steers received a high antagonist growing diet (analyzed 4.9 mg Cu/kg DM, 0.48% S, and 5.3 mg Mo/kg DM). Steers were weighed at the beginning (days 1 and 0) and end (days 125 and 126) of the trial to determine average daily gain (ADG) and gain:feed (G:F). Blood was collected from all steers on days 0, 28, 56, 84, and 126. Liver samples were collected on days −3 or −2 and day 123 or 124. Data were analyzed using ProcMixed of SAS (experimental unit = steer; fixed effect = treatment and block). Plasma Cu was analyzed as repeated measures (repeated effect = day). Plasma and liver Cu concentrations were regressed against total Cu intake using ProcGLM to calculate relative bioavailability of GLY. Final BW and overall ADG were greatest for CS5 and CS10 and least for CON and GLY5 (P = 0.01). Overall, DMI was not affected by treatment (P = 0.14), but overall G:F tended to be greatest for CS5, CS10, and GLY5 and least for CON (P = 0.08). Total and supplemental Cu intake was greatest for steers supplemented either source at 10 mg Cu/kg DM and least for CON (P < 0.01). However, total and supplemental Cu intake was greater for CS5 than GLY5 (P < 0.01). Final liver Cu concentrations were greatest for CS10, least for CON, CS5, and CS10, and intermediate for GLY10 (P < 0.01). Final plasma Cu was greatest for steers supplemented either source at 10 mg Cu/kg DM (P < 0.01). Relative bioavailability of GLY was 82% compared to CS based on liver Cu (P < 0.01) but did not differ based on plasma Cu (P = 0.60). The lesser bioavailability of GLY relative to CS could be due to a high concentration of dietary antagonists and lower solubility of GLY (68.9% relative to CS) in pH conditions (5.2) similar to the ruminal pH of a beef animal consuming a high concentrate diet. Future studies should examine the effects of bis-glycinate bound Cu fed in blended combination with inorganic Cu sulfate to determine the most effective blend of sources for feedlot cattle experiencing varying amounts of dietary Cu antagonists.     

Validating an empiric sulfadiazine–trimethoprim dosage regimen for treatment of Escherichia coli and Staphylococcus delphini infections in mink (Neovison vison)

Antimicrobial agents are used extensively off‐label in mink, as almost no agents are registered for this animal species. Pharmacokinetic (PK) and pharmacodynamic (PD) data are required to determine antimicrobial dosages specifically targeting mink bacterial pathogens. The aims of this study were to assess, in a PKPD framework, the empirical dosage regimen for a combination of trimethoprim (TMP) and sulfadiazine (SDZ) in mink, and secondarily to produce data for future setting of clinical breakpoints. TMP and SDZ PK parameters were obtained experimentally in 22 minks following IV or oral administration of TMP/SDZ (30 mg/kg, i.e. 5 mg/kg TMP and 25 mg/kg SDZ). fAUC/MIC with a target value of 24 hr was selected as the PKPD index predictive of TMP/SDZ efficacy. Using a modeling approach, PKPD cutoffs for TMP and SDZ were determined as 0.062 and 16 mg/L, respectively. By incorporating an anticipated potentiation effect of SDZ on TMP against Escherichia coli and Staphylococcus delphini, the PKPD cutoff of TMP was revised to 0.312 mg/L, which is above the tentative epidemiological cutoffs (TECOFF) for these species. The current empirical TMP/SDZ dosage regimen (30 mg/kg, PO, once daily) therefore appears adequate for treatment of wild‐type E. coli and S. delphini infections in mink.     

Key Factors Affecting Reproductive Success of Thoroughbred Mares and Stallions on a Commercial Stud Farm

To evaluate factors contributing to fertility of thoroughbred mares, data from 3743 oestrous periods of 2385 mares were collected on a large thoroughbred farm in Ireland. Fourteen stallions (mean age 8.3 years; range 4–15 years) had bred 2385 mares (mean age 9.4 years; range 3–24 years). Maiden mares accounted for 12%, mares with a foal at foot for 64%, and barren, slipped or rested mares for 24% of the total. The mean pregnancy rate per cycle was 67.8% (68.6% in year 1 and 66.9% in year 2). Backward stepwise multivariable logistic regression analysis was utilized to develop two models to evaluate mare factors, including mare age, reproductive status, month of foaling, dystocia, month of cover, foal heat, cycle number, treatments, walk‐in status and stallion factors including stallion identity, stallion age, shuttle status, time elapsed between covers and high stallion usage on the per cycle pregnancy rate and pregnancy loss. Old age (p < 0.001) and cover within 20 days post‐partum (p < 0.003) were associated with lowered pregnancy rates. High mare age (p < 0.05) and barren, slipped or rested reproductive status (p = 0.05) increased the likelihood of pregnancy loss. Uterine inflammation or infection, if appropriately treated, did not affect fertility. Only high usage of stallions (used more than 21 times in previous week) was associated with lowered (p = 0.009) pregnancy rates. However, shuttle stallions were more likely to have increased (p = 0.035) pregnancy survival, perhaps reflecting a bias in stallion selection. In conclusion, mare age exerted the greatest influence on fertility; nonetheless, thoroughbreds can be effectively managed to achieve high reproductive performance in a commercial setting.