Evaluation of the specificity of three enzyme-linked immunosorbent assays for detection of antibodies against Salmonella in bovine bulk milk

Background

The Swedish Salmonella control program has been running for decades and has resulted in a low prevalence of Salmonella in Swedish food producing animals. Routine bacteriology is used to detect Salmonella, however, bacteriology is time consuming, costly and has a low sensitivity. Different enzyme-linked immunosorbent assays (ELISAs) have been developed for detection of antibodies against Salmonella Dublin and S. Typhimurium in bovine bulk milk, individual milk samples as well as in sera. Screening bulk milk for antibodies against Salmonella spp. could improve the cost-effectiveness of the surveillance in Swedish dairy cattle, but as characteristics of tests may vary in different populations, tests should always be evaluated in the specific population where they will be used. Hence, the aim of this study was to evaluate the specificities of three bovine ELISAs when used to analyse bulk milk samples from Swedish dairy cattle. A second aim was to compare the performance of the two Dublin ELISAs tested.

Methods

Bulk milk samples for analysis were randomly selected from samples collected within the Swedish bulk milk sampling scheme and analyzed with the three ELISAs; a Danish in-house Dublin ELISA, PrioCHECK® Salmonella Ab bovine Dublin ELISA and PrioCHECK® Salmonella Ab bovine ELISA (hereafter named mixed ELISA). The specificities of the ELISAs were calculated assuming a disease-free status in Sweden i.e. that all test positive samples were assumed to be false positive results. This assumption can be used when a disease is known to be infrequent.

Results

The calculated specificities of the two Dublin ELISAs and the mixed ELISA, when using the producer’s recommended cut-off value of the corrected optic-density percent (ODC%) were 99.4% (95% Confidence Interval (CI): 98.8% -99.8%), 99.4% (95% CI: 98.8% -99.8%) and 97.9% (95% CI: 96.8% -98.7%), respectively. The correlation between the ODC% values of the two Dublin ELISAs was 0.83.

Conclusions

We conclude that the evaluated ELISAs have sufficiently high specificities to be used as supplement to bacteriological examinations in the Swedish Salmonella control program in cattle as well as a primary screening test in routine surveillance for S. Dublin.     

Effect of dry period length on the effect of an intramammary teat sealant on the risk of mastitis in cattle treated with antibiotics at drying off

AIM: The aim of this study was to evaluate, under farm conditions, the use of a teat sealant in addition to whole herd dry cow antibiotic therapy on the risk of clinical mastitis in dairy cattle at pasture, and to evaluate the impact of dry period length on that risk and the impact of the teat sealant on that risk.

METHODS: Dairy cows in three herds which used routine whole herd antibiotic therapy were randomly assigned to receive either treatment with an internal teat sealant (n=322) or no additional treatment (n=313) at drying-off between March and May 2010. All clinical mastitis cases during the dry period and to the end of the subsequent lactation were recorded by farm staff; factors affecting risk of clinical mastitis were then analysed using a Cox proportional hazards model.

RESULTS: Median duration of the dry period was 112 days with >25% of cows having a dry period >130 days. The incidence risk of mastitis during lactation for cows treated with teat sealant was 9.9 (95% CI=6.9–13.7) cases per 100 cows compared with 17.9 (95% CI=13.8–22.6) cases per 100 cows for cows treated with antibiotic alone. The addition of a teat sealant to dry cow antibiotic therapy decreased the risk of clinical mastitis only in the first 33 days after calving (Hazard risk 0.24 (95% CI=0.12–0.48)). Length of dry period did not significantly affect the risk of clinical mastitis, or the effect of adding teat sealant to dry cow antibiotic therapy on the risk of clinical mastitis.

CONCLUSIONS: In these herds where, based on the mastitis history, whole herd antibiotic therapy had been recommended, the use of a teat sealant significantly reduced the risk of clinical mastitis. This effect was limited to the first 33 days after calving; subsequently there was no significant effect of treatment. There was no effect of dry period length on risk of clinical mastitis, nor any significant interaction with treatment.

CLINICAL RELEVANCE: Combination therapy with teat sealant and antibiotic was effective under New Zealand conditions in herds using whole herd antibiotic treatment at drying off. Teat sealant reduced risk of clinical mastitis in cattle with dry periods substantially longer than 100 days, and there was no evidence that this effect changed as dry period length increased.     

When less is more: heterogeneity in grass patch height supports herbivores in counter-intuitive ways

Herbivores are an integral part of the African landscape and have evolved with the vegetation to create the savanna landscape. Managers of these landscapes can benefit from a better understanding of how indigenous herbivores use the landscape to which they are adapted. In this study we observed which patches were frequently utilised, by doing regular monthly road counts, grass height observations and dung counts on selected short grass patches in the Kruger National Park. Smaller-framed impala and blue wildebeest (meso-herbivores) were most regularly seen on these nutritious patches, while from dung deposits it was clear that the even larger-framed buffalo (mega-herbivores) spent time there. This preference can be explained by considering the nutritional needs and food intake of the herbivores. Smaller-framed herbivores seem to be able to satisfy their dietary requirements on the high-quality forage patches, while larger-framed herbivores seem to supplement the quality forage by also spending foraging time on areas of higher grass biomass. From this insight we propose that range management should take herbivore preferences into account and allow herbivores to select and concentrate their foraging on the most nutritious forage. This approach is likely to decrease inputs while allowing animals to maintain or increase production.     

Preliminary pharmacokinetics of tramadol hydrochloride after administration via different routes in male and female B6 mice

Objective

1) To determine the pharmacokinetics of tramadol hydrochloride and its active metabolite, O-desmethyltramadol (M1), after administration through different routes in female and male C57Bl/6 mice; 2) to evaluate the stability of tramadol solutions; and 3) to identify a suitable dose regimen for prospective clinical analgesia in B6 mice.

Bone Morphogenetic Protein‐6 (BMP‐6) Stimulates the Antrum Formation by the Regulation of its Signalling Pathway in Caprine Pre‐antral Follicles Cultured In Vitro

BMP‐6 has been found to be important to ovarian cells and oocyte, as well as to uterus. Thus, this study investigated the effect of bone morphogenetic protein (BMP‐6) and recombinant follicle‐stimulating hormone (rFSH) alone or in combination on the in vitro culture (IVC) of isolated caprine secondary follicles (Experiment 1) and the mRNA levels for BMP receptors/Smad signalling pathway (BMPR1A, BMPR2, SMAD1, SMAD4, SMAD5, SMAD6, SMAD7 and SMAD8) in vivo and in vitro using BMP‐6 (Experiment 2). Secondary follicles were cultured in αMEM⁺ alone (control medium) or supplemented with BMP‐6 at 1 or 10 ng/ml and rFSH alone or the combination of both BMP‐6 concentrations and rFSH. The results from Experiment 1 showed that the antrum formation rate was higher in the BMP‐6 at 1 ng/ml (p < 0.05) than in MEM. In Experiment 2, the mRNA expression for BMPR2, SMAD1, SMAD5 and SMAD6 was detected in non‐cultured control and after in vitro culture (MEM and 1 ng/ml BMP‐6); while the expression of SMAD7 and SMAD8 mRNA was only detected after IVC, SMAD4 was only detected in the BMP‐6 at 1 ng/ml treatment. In conclusion, the low BMP‐6 concentration positively influenced antrum formation and ensured normal mRNA expression for BMP receptor and Smads after IVC of caprine secondary follicles.     

Epigallocatechin‐3‐gallate (EGCG) and green tea polyphenols do not improve stallion semen parameters during cooling at 4°C

Stallion semen storage for artificial insemination is mainly based on liquid cooled storage. In many stallions this technique maintains sperm quality for an extended period of time (24–72 hr) at 7°C. While this technique is commonly used in the horse industry, there can be a decline in fertility in some stallions, due to an inability of their sperm to tolerate the cool storage process. The aim of the present work was to evaluate the effect of two natural antioxidants (epigallocatechin‐3‐gallate (EGCG) at 20, 60 and 120 μm and green tea polyphenols, and p at .001, .01 and .1 mg/ml) on some sperm parameters (sperm motility, viability/acrosome integrity and DNA quality) in extended semen immediately after its collection (T0) and after 2, 6, 24 and 48 hr of cool storage. Two ejaculates from three trotter stallions were analysed after 48 hr of storage at 4°C. No beneficial effect on the analysed parameters was observed: the two antioxidants were not able to improve sperm quality after 48 hr of storage. These results are in agreement with previous findings on the effect of different antioxidants reported by other researches, who have demonstrated that stallion semen keeps good antioxidant capacity after dilution for 24 hr. In conclusion, the positive effect exerted by antioxidant molecules in other species is not confirmed in the equine one.     

Detection of intracellular reactive oxygen species (superoxide anion and hydrogen peroxide) and lipid peroxidation during cryopreservation of alpaca spermatozoa

The objective of this study was to detect changes in intracellular reactive oxygen species (superoxide anion and hydrogen peroxide) production and lipid peroxidation during cryopreservation of alpaca spermatozoa. Twelve alpaca semen samples were conventionally cryopreserved. Intracellular superoxide anion and hydrogen peroxide were evaluated by fluorescence microscopy using dihydroethidium (DHE)/YO‐PRO‐1 and dichlorofluorescein diacetate (H2DCFDA)/propidium iodide (PI), respectively. Evaluations were performed during cooling curve at (1) 25°C, (2) 15°C, (3) 5°C/0 min, (4) 5°C/15 min, (5) 5°C/30 min and (6) after freezing/thawing. Evaluation of lipid peroxidation by measuring malondialdehyde (MDA) was performed at 25°C, 5°C/30 min and after thawing. Maximum percentages of total spermatozoa producing superoxide anion and hydrogen peroxide were found at 5°C/30 min (62.8 ± 6.3% and 30.5 ± 5.6%, respectively), and these results were higher (p < .05) than initial (25°C: 10.8 ± 3.8% and 6.8 ± 0.7%, respectively) and after thawing (29.8 ± 9.5% and 7.5 ± 1.8%, respectively) values. However, considering only viable spermatozoa, production of superoxide anion and hydrogen peroxide during overall stabilization at 5°C (>76% and >91%, respectively) and after thawing (74.9 ± 5.0% and 78.9 ± 2.2%, respectively) was higher (p < .05) than initial values at 25°C (38.7 ± 3.1% and 53.6 ± 2.0%, respectively). Lipid peroxidation at 25°C, 5°C/30 min, and post‐thawing were 346.5 ± 99.8, 401.1 ± 64.8 and 527.7 ± 142.8 ng/ml MDA, respectively. These results showed that high percentage of viable alpaca spermatozoa produces intracellular reactive species oxygen (ROS) during the cryopreservation process of alpaca semen.     

Variability of the nitrifying bacteria in the biofilm and water column of a recirculating aquaculture system for tilapia (Oreochromis niloticus) production

The aim of this study was to evaluate variability of nitrifying bacterial community in the biofilm and in the water of a recirculating aquaculture systems (RAS) in a tilapia farming in order to determine if nitrification process is dependent, or not, of nitrifying bacteria abundance. Biofilm and water samples were collected periodically for 30 days and analysed with the fluorescent in situ hybridization (FISH) technique, used to quantify ammonia‐oxidizing bacteria (AOB) and nitrite‐oxidizing bacteria (NOB). Ammonia presented the peak in the first week, while the nitrite's maximum was recorded in the second week. Nitrate increased steadily, indicating nitrification activity. Total bacterial abundance in biofilm increased continuously, while in water, it did not change significantly. In the biofilm, number of AOB was high at beginning, decreased after few days and increased again following augment of ammonia. Number of NOB also showed an increase in abundance in biofilm following the increment of nitrite and nitrate. In water, AOB and NOB did not show major variability. Relative abundance of nitrifying bacteria represented more than 30% of total bacteria in biofilm at beginning of the experiment. Their contribution decreased to >3% in last days. It indicates that nitrifying bacteria are biofilm colonizers, and that their activity seems to be directly related to the concentration of nitrogen compounds. However, contribution of nitrifying bacteria did not vary much along the time. We may conclude that the biofilm‐nitrifying bacteria plays major role in nitrification process in RAS and that the activity of these organisms is dependent of their abundance in response to the concentration of nitrogen compounds.     

Seasonal dynamics of microbial community in an aquaculture system for Nile tilapia (Oreochromis niloticus)

Studies on bacterial abundance and diversity will improve the understanding of the microbial ecology to optimize aquaculture production, water quality, disease control and environmental impact of effluents. We comparatively evaluated an aquaculture system in dry and rainy seasons by Fluorescent in situ Hybridization (FISH) and culture‐dependent methodology. Overall, a negative correlation between seasonality in bacterial and flagellates abundance was observed. Alpha‐, beta‐, gamma‐proteobacteria and Cytophaga‐Flavobacter were observed with seasonal variation. Putative pathogenic bacteria such as coagulase‐negative Staphylococcus, non‐fermenter Gram‐negative rods (Pseudomonas sp. and Burkholderia sp.), and Enterobacteriaceae were also detected in this study with significant seasonal variation. Focusing on medically important bacteria, our data show that microbial diversity in the environment associated with aquaculture, as it is practiced, may be altered in growth ponds used for fish cultivation. As an ecological consequence, potentially pathogenic bacteria might be released in high concentrations to the downstream environments posing potential threats to human and animal health.     

Potential of Community-Based Forest Management to Mitigate Climate Change in the Philippines

Community-based forest management (CBFM) is the principal strategy in managing inhabited ‘forest land’ in the Philippines. It involves the participation of local communities in various forestry activities to achieve sustainable forestry, advance social justice and improve socioeconomic welfare, and promote a stable and healthy environment. This paper analyses the potential benefits of agroforestry farms in CBFM sites to mitigate climate change. The incorporation of trees in farms and landscapes has led to enhanced carbon storage and sequestration. Half a million hectares of agroforestry farms in CBFM sites in the Philippines are estimated to store 25 MtC while sequestering 2.7 MtC annually. Lessons are drawn from three carbon sequestration projects under development using CBFM as the main approach. Income from carbon credits is not sufficient to recover the cost of tree planting. The transaction costs of forestry CDM projects are substantial and could prove to be the greatest barrier to project fruition. Government institutions must find ways to encourage project developers by simplifying rules and regulations for forestry carbon projects. Forest definition must be assessed. Project developers and the government could also explore the voluntary carbon market which is more flexible than the CDM market. Policy and technical studies must be conducted to ascertain the potential of the REDD for CBFM sites in the country.