Physicochemical properties of micellar casein retentates generated at different microfiltration temperatures.

Processing temperature has a significant influence on the composition and functionality of the resulting streams following microfiltration (MF) of skim milk. In this study, MF and diafiltration (DF) were performed at 4 or 50°C to produce ß-casein (ß-CN)-depleted and nondepleted (i.e., native casein profile) micellar casein isolate retentates, respectively. Microfiltration combined with extensive DF resulted in a 40% depletion of ß-CN at 4°C, whereas no ß-CN depletion occurred at 50°C. Microfiltration at 4°C led to higher transmission of calcium into permeates, with retentate generated at 4°C containing less total calcium compared with retentate generated at 50°C, based on the volume of retentate remaining. Higher heat stability at 120°C was measured for retentates generated at 4°C compared with those at 50°C, across all pH values measured. Retentates generated at 4°C also had significantly lower ionic calcium values at each pH compared with those generated at 50°C. Higher apparent viscosities at 4°C were measured for retentates generated at 4°C compared with retentates generated at 50°C, likely due to increased voluminosity of ß-CN-depleted casein micelles. The results of this study provide new information on how changing the composition of MF retentate, by appropriate control of processing temperature and DF, can alter physicochemical properties of casein micelles, with potential implications for ingredient functionality.

Effect of reverse osmosis and ultra-high-pressure homogenization on the composition and microstructure of sweet buttermilk.

Buttermilk (BM), the by-product of butter making, is similar to skim milk (SM) composition. However, it is currently undervalued in dairy processing because it is responsible for texture defects (e.g., crumbliness, decreased firmness) in cheese and yogurt. One possible way of improving the incorporation of BM into dairy products is by the use of technological pretreatments such as membrane filtration and homogenization. The study aimed at characterizing the effect of preconcentration by reverse osmosis (RO) and single-pass ultra-high-pressure homogenization (UHPH) on the composition and microstructure of sweet BM to modify its techno-functional properties (e.g., protein gel formation, syneresis, firmness). The BM and RO BM were treated at 0, 15, 150, and 300 MPa. Pressure-treated and control BM and RO BM were ultracentrifuged to fractionate them into the following 3 fractions: a supernatant soluble fraction (top layer), a colloidal fraction consisting of a cloudy layer (middle layer), and a high-density pellet (bottom layer). Compositional changes in the soluble fraction [lipid, phospholipid (PL), protein, and salt], as well as its protein profile by PAGE analysis, were determined. Modifications in particle size distribution upon UHPH were monitored by laser diffraction in the presence and absence of sodium citrate to dissociate the casein (CN) micelles. Microstructural changes in pressure-treated and non-pressure-treated BM and RO BM particles were monitored by confocal laser scanning microscopy. Particle size analysis showed that UHPH treatment significantly decreased the size of the milk fat globule membrane fragments in BM and RO BM. Also, pressure treatment at 300 MPa led to a significant increase in the recovery of total lipids, CN, calcium, and phosphate in the BM soluble fraction (top layer) following ultracentrifugation. However, PL were primarily concentrated in the pellet cloud (middle layer), located above the pellet in BM concentrated by RO. In contrast, PL were evenly distributed between soluble and colloidal phases of BM. This study provides insight into the modifications of sweet BM constituents induced by RO and UHPH from a compositional and structural perspective.

Production and storage stability of concentrated micellar casein.

Concentrated micellar casein (CMC) is a high-protein ingredient that can be used in process cheese product formulations. The objectives of this study were to develop a process to produce CMC and to evaluate the effect of sodium chloride and sodium citrate on its storage stability. Skim milk was pasteurized at 76°C for 16 s and cooled to ≤4°C. The skim milk was heated to 50°C using a plate heat exchanger and microfiltered with a graded permeability (GP) ceramic microfiltration (MF) membrane system (0.1 µm) in a continuous feed-and-bleed mode (flux of 71.43 L/m2 per hour) using a 3× concentration factor (CF) to produce a 3× MF retentate. Subsequently, the retentate of the first stage was diluted 2× with soft water (2 kg of water: 1 kg of retentate) and again MF at 50°C using a 3× CF. The retentate of the second stage was then cooled to 4°C and stored overnight. The following day, the retentate was heated to 63°C and MF in a recirculation mode until the total solids (TS) reached approximately 22% (wt/wt). Subsequently, the MF system temperature was increased to 74°C and MF until the permeate flux was <3 L/m2 per hour. The CMC was then divided into 3 aliquots (approximately 10 kg each) at 74°C. The first portion was a control, whereas 1% of sodium chloride was added to the second portion (T1), and 1% of sodium chloride plus 1% of sodium citrate were added to the third portion (T2). The CMC retentates were transferred hot to sterilized vials and stored at 4°C. This trial was repeated 3 times using separate lots of skim milk. The CMC at d 0 (immediately after manufacturing) contained 25.41% TS, 21.65% true protein (TP), 0.09% nonprotein nitrogen (NPN), and 0.55% noncasein nitrogen (NCN). Mean total aerobic bacterial counts (TBC) in control, T1, and T2 at d 0 were 2.6, 2.5, and 2.8 log cfu/mL, respectively. The level of proteolysis (NCN and NPN values) increased with increasing TBC during 60 d of storage at 4°C. This study determined that CMC with >25% TS and >95% casein as percentage of TP can be manufactured using GP MF ceramic membranes and could be stored up to 60 d at 4°C. The effects of the small increase in NCN and NPN, as well as the addition of sodium chloride or sodium citrate in CMC during 60 d of storage on process cheese characteristics, will be evaluated in subsequent studies.

Forward osmosis concentration of milk: Product quality and processing considerations.

Concentration of milk in the dairy industry is typically achieved by thermal evaporation or reverse osmosis (RO). Heat concentration is energy intensive and leads to cooked flavor and color changes in the final product, and RO is affected by fouling, which limits the final achievable concentration of the product. The main objective of this work was to evaluate forward osmosis (FO) as an alternative method for concentrating milk. The effects of fat content and temperature on the process were evaluated, and the physicochemical properties and sensory qualities of the final product were assessed. Commercially pasteurized skim and whole milk samples were concentrated at 4, 15, and 25°C using a benchtop FO unit. The FO process was assessed by monitoring water flux and product concentration. The color of the milk concentrates was also evaluated. A sensory panel compared the FO concentrated and thermally concentrated milks, diluted to single strength, with high temperature, short time pasteurized milk. The FO experimental runs were conducted in triplicate, and data were analyzed by single-factor ANOVA. Water flux during FO decreased with time under all processing conditions. Higher temperatures led to faster concentration and higher concentration factors for both skim and whole milk. After 5.75 h of FO processing, the concentration factors achieved for skim milk were 2.68 ± 0.08 at 25°C, 2.68 ± 0.09 at 15°C, and 2.36 ± 0.08 at 4°C. For whole milk, after 5.75 h of FO processing, concentration factors of 2.32 ± 0.12 at 25°C, 2.12 ± 0.36 at 15°C, and 1.91 ± 0.15 at 4°C were obtained. Overall, maximum concentration levels of 40.15% total solids for skim milk and 40.94% total solids for whole milk were achieved. Additionally, a triangle sensory test showed no significant differences between regular milk and FO concentrated milk diluted to single strength. This work shows that FO is a viable nonthermal processing method for concentrating milk, but some technical challenges need to be overcome to facilitate commercial utilization.

Efficiency of removal of whey protein from sweet whey using polymeric microfiltration membranes.

Our objective was to measure whey protein removal percentage from separated sweet whey using spiral-wound (SW) polymeric microfiltration (MF) membranes using a 3-stage, 3× process at 50°C and to compare the performance of polymeric membranes with ceramic membranes. Pasteurized, separated Cheddar cheese whey (1,080 kg) was microfiltered using a polymeric 0.3-µm polyvinylidene (PVDF) fluoride SW membrane and a 3×, 3-stage MF process. Cheese making and whey processing were replicated 3 times. There was no detectable level of lactoferrin and no intact α- or ß-casein detected in the MF permeate from the 0.3-µm SW PVDF membranes used in this study. We found BSA and IgG in both the retentate and permeate. The ß-lactoglobulin (ß-LG) and α-lactalbumin (α-LA) partitioned between retentate and permeate, but ß-LG passage through the membrane was retarded more than α-LA because the ratio of ß-LG to α-LA was higher in the MF retentate than either in the sweet whey feed or the MF permeate. About 69% of the crude protein present in the pasteurized separated sweet whey was removed using a 3×, 3-stage, 0.3-µm SW PVDF MF process at 50°C compared with 0.1-µm ceramic graded permeability MF that removed about 85% of crude protein from sweet whey. The polymeric SW membranes used in this study achieve approximately 20% lower yield of whey protein isolate (WPI) and a 50% higher yield of whey protein phospholipid concentrate (WPPC) under the same MF processing conditions as ceramic MF membranes used in the comparison study. Total gross revenue from the sale of WPI plus WPPC produced with polymeric versus ceramic membranes is influenced by both the absolute market price for each product and the ratio of market price of these 2 products. The combination of the market price of WPPC versus WPI and the influence of difference in yield of WPPC and WPI produced with polymeric versus ceramic membranes yielded a price ratio of WPPC versus WPI of 0.556 as the cross over point that determined which membrane type achieves higher total gross revenue return from production of these 2 products from separated sweet whey. A complete economic engineering study comparison of the WPI and WPPC manufacturing costs for polymeric versus ceramic MF membranes is needed to determine the effect of membrane material selection on long-term processing costs, which will affect net revenue and profit when the same quantity of sweet whey is processed under various market price conditions.

Determination of the efficiency of removal of whey protein from sweet whey with ceramic microfiltration membranes.

Our research objective was to measure percent removal of whey protein from separated sweet whey using 0.1-µm uniform transmembrane pressure ceramic microfiltration (MF) membranes in a sequential batch 3-stage, 3× process at 50°C. Cheddar cheese whey was centrifugally separated to remove fat at 72°C and pasteurized (72°C for 15 s), cooled to 4°C, and held overnight. Separated whey (375 kg) was heated to 50°C with a plate heat exchanger and microfiltered using a pilot-scale ceramic 0.1-µm uniform transmembrane pressure MF system in bleed-and-feed mode at 50°C in a sequential batch 3-stage (2 diafiltration stages) process to produce a 3× MF retentate and MF permeate. Feed, retentate, and permeate samples were analyzed for total nitrogen, noncasein nitrogen, and nonprotein nitrogen using the Kjeldahl method. Sodium dodecyl sulfate-PAGE analysis was also performed on the whey feeds, retentates, and permeates from each stage. A flux of 54 kg/m2 per hour was achieved with 0.1-µm ceramic uniform transmembrane pressure microfiltration membranes at 50°C. About 85% of the total nitrogen in the whey feed passed though the membrane into the permeate. No passage of lactoferrin from the sweet whey feed of the MF into the MF permeate was detected. There was some passage of IgG, bovine serum albumen, glycomacropeptide, and casein proteolysis products into the permeate. ß-Lactoglobulin was in higher concentration in the retentate than the permeate, indicating that it was partially blocked from passage through the ceramic MF membrane.

Efficiency of three boar sperm enrichment techniques.

The objective of the study was to investigate the efficiency of three enrichment methods to separate boar spermatozoa. Twenty-four ejaculates from 12 boars (2 ejaculates/boar) were extended (30 × 106 spermatozoa/mL) in commercial Beltsville Thawing Solution. Each semen sample was processed with glass wool column (GW) and glass beads (GB) filtration and with the single-layer centrifugation (SLC) technique. Semen samples before (control; C) and after treatment were evaluated for sperm CASA motility/kinetics and concentration, viability, morphology and chromatin integrity. Data were analysed with mixed models. The concentration of total and motile spermatozoa was significantly decreased after treatment in groups GW and SLC, but not in group GB. Group GW showed increased values of WOB compared with both groups C and GB. Group GB showed greater values of rapid movement spermatozoa and lower values of slow movement spermatozoa compared with group C. In group SLC, higher values of VSL, LIN and STR were observed compared with group C. In conclusion, all techniques under examination enhanced various CASA variables. Based on our results, the GB method is a promising alternative separation technique for boar sperm and deserves further research regarding swine in vitro fertilization.

Bovine colostrum whey: Postpartum changes of particle size distribution and immunoglobulin G concentration at different filtration pore sizes.

Bovine colostrum, as vital as it is for calves, is also a valuable source of functional components with rich health benefits for humans. Bovine colostrum whey consists of a large number of bioactive proteins and peptides. The most abundant of these is IgG. Particle size distribution (PSD) is an important feature of many of the processes in the dairy food industries. Despite this, scientific literature on PSD of colostrum whey is scarce. The goal of this research was to describe bovine colostrum whey PSD with an emphasis on postpartum milking time, filtration (pore size 450, 100, and 20 nm), IgG concentration, and lactation number. For this purpose, 4 postpartum milking colostrum samples were sequentially milked from 46 Holstein cows at 12 ± 1 h intervals. Colostrum whey was prepared by renneting and diluted (1:200) for PSD analyses by a Malvern Zetasizer Nano ZS (Malvern Instruments Ltd., Malvern, UK). Immunoglobulin G concentration of these diluted colostrum whey samples were analyzed by an Octet K2 (Molecular Devices LLC, San Jose, CA) system. Linear mixed model analysis revealed significant effects of filter pore size, postpartum milking, and lactation on colostrum whey IgG concentrations. The percentage of particles in the size interval 5 to 15 nm (the hydrodynamic diameter of IgG is around 10 nm) had an intermediate positive correlation (r = 0.50) with IgG concentration. Furthermore, we showed that PSD was associated with IgG concentration, postpartum milking time, and lactation number. The PSD measurement results showed the mean hydrodynamic diameter of 100 nm pore size filtered colostrum whey to be around 10 nm. This, with the IgG concentration results, suggests that even though the size of IgG is around 10 nm, a 100 nm pore size is adequate for membrane-involved IgG separations. In terms of energy efficiency of the filtration process, the use of a larger filter pore size can make a remarkable difference, for example, in pressurizing and cooling costs. Our work contributes to the development of sustainable and widely available colostrum-derived food and feed supplements.

Influence of a cell salvage washing system and leukocyte reduction filtration on bacterial contamination of canine whole blood ex vivo.

OBJECTIVE: To determine the ability of cell salvage washing and leukoreduction filtration to remove bacterial contamination from canine whole blood. STUDY DESIGN: Ex vivo nested cohort study. SAMPLE POPULATION: Commercially purchased fresh canine whole blood (n = 33 units). METHODS: Commercially obtained canine whole blood was inoculated with known concentrations of one of three species of bacteria, Escherichia coli (ATCC 25922), Staphylococcus pseudintermedius (quality control strain; Texas A&M University), or Pseudomonas aeruginosa (ATCC 27853). Negative controls were inoculated with sterile saline. The inoculated blood was processed through a cell salvage system and filtered through a series of two leukocyte reduction filters. Samples were aseptically collected at five points during processing (inoculum, prewash, postwash, post-first filtration, and post-second filtration) for bacterial enumeration. RESULTS: Bacterial concentrations were reduced by 85.2%, 91.5%, and 93.9% for E coli, S pseudintermedius, and P aeruginosa, respectively, after washing (P < .0001), and bacterial concentrations were reduced by 99.9%, 100%, and 100%, respectively, after the first filtration (P < .0001). After the second filtration, none of the three species of bacteria could be isolated (100% reduction). No bacterial growth was obtained from negative controls throughout the study. The type of bacteria (P = .29) did not allow prediction of bacterial reduction. CONCLUSION: Cell salvage washing combined with leukoreduction filtration eliminated bacterial contamination of whole dog blood (P < .0001). CLINICAL SIGNIFICANCE: Cell salvage washing and leukoreduction filtration could be applied to intraoperative autotransfusion in clinical animals, especially those treated for trauma or hemorrhage with concurrent bacterial contamination.

Evaluation of a filter system to harvest plasma for identification of failure of passive transfer in newborn calves.

The objective of this study was to evaluate a filter system to harvest plasma to assess failure of passive transfer (FPT) in newborn calves. Blood samples (n = 227) for serum and plasma harvesting were collected via jugular vein puncture from Holstein calves aged 1 to 7 d from 4 commercial dairy herds in Northeast Germany. Serum IgG concentrations were determined using a sandwich ELISA. Failure of passive transfer was defined as IgG concentrations <10 mg/mL and used as a gold standard. One handheld optical refractometer (Euromex Holland, Arnhem, the Netherlands) and 2 digital Brix refractometers (device 1: HI 96801 digital refractometer, Hanna Instruments, Woonsocket, RI; device 2: Misco PA201, Misco, Solon, OH) were used to analyze total proteins in serum or plasma. The colostrum uptake of the calf can thus be monitored and calves with FPT can be identified. Serum was obtained through centrifugation. Plasma was obtained through either a filter system or centrifugation. For plasma filtration, approximately 2 mL of lithium heparin blood was injected into the inlet reservoir of a plasma filter (2-Drop-Filter, Pharmadoc, Lübeck, Germany) using a disposable syringe. Receiver operating characteristic curve analyses were used to determine optimum thresholds for each of the 3 devices using different media. Sixty-seven (30%) calves had FPT. For the handheld optical refractometer, the optimum threshold was 5.6 g/dL [sensitivity 70.1%; specificity 80.0%; positive predictive value (PPV) 60.1%; negative predictive value (NPV) 86.2%; area under the curve (AUC) 0.85] using serum. For centrifuged plasma, the optimum threshold was 6.3 g/dL (sensitivity 82.1%; specificity 68.1%; PPV 52.5%; NPV 89.9%; AUC 0.84), and for filtered plasma, the threshold was 6.0 g/dL (sensitivity 56.7%; specificity 90.0%; PPV 70.9%; NPV 82.9%; AUC 0.80). For device 1, the optimum threshold was 8.9% Brix (sensitivity 82.1%; specificity 63.8%; PPV 48.7%; NPV 89.5%; AUC 0.81), 9.4% Brix (sensitivity 76.1%; specificity 73.7%; PPV 55.4%; NPV 87.8%; AUC 0.80), using serum and centrifuged plasma, respectively. For device 2, the optimum threshold was 8.7% Brix (sensitivity 74.6%; specificity 76.2%; PPV 57.4%; NPV 87.5%; AUC 0.83), 9.5% Brix (sensitivity 80.6%; specificity 70.6%; PPV 54.0%; NPV 89.5%; AUC 0.83), and 9.2% Brix (sensitivity 58.2%; specificity 87.5%; PPV 66.6%; NPV 83.0%; AUC 0.80) using serum, centrifuged plasma, and filtered plasma, respectively. Based on the AUC, the 3 devices yielded comparable test characteristics to identify calves with FPT. In conclusion, a filter system can be used to facilitate the evaluation of FPT as a point of care technique in calves without the need for serum centrifugation.

A comparison of the heat stability of fresh milk protein concentrates obtained by microfiltration, ultrafiltration and diafiltration.

The objective of this work was to evaluate the impact of changes during membrane filtration on the heat stability of milk protein concentrates. Dairy protein concentrates have been widely employed in high protein drinks formulations and their stability to heat treatment is critical to ensure quality of the final product. Pasteurized milk was concentrated three-fold by membrane filtration, and the ionic composition was modified by addition of water or permeate from filtration (diafiltration). Diafiltration with water did not affect the apparent diameter of the casein micelles, but had a positive effect on heat coagulation time (HCT), which was significantly longer (50 min), compared to the non diafiltered concentrates (about 30 min). UHT treatments increased the particle size of the casein micelles, as well as the turbidity of retentates. Differences between samples with and without diafiltration were confirmed throughout further analysis of the protein composition of the unsedimentable fraction, highlighting the importance of soluble protein composition on the processing functionality of milk concentrates.

Sperm preparation through Sephadex™ filtration improves in vitro fertilization rate of buffalo oocytes.

Routinely, swim-up method is used to separate high-quality sperm; however, long processing time and close cell-to-cell contact during the centrifugation step are inevitable elements of oxidative stress to sperm. The objective was to evaluate Sephadex™ and glass wool filtration to separate motile, intact and viable sperm for in vitro fertilization in buffalo. The cumulus-oocyte complexes (COCs) were collected from ovaries of slaughtered buffaloes by aspiration and matured for 24 hr in CO2 incubator at 38.5°C and 5% CO2 . Matured COCs were rinsed twice in fertilization TALP and placed in the pre-warmed fertilization medium without sperm. Cryopreserved buffalo semen was thawed at 37°C for 30 s and processed through Sephadex™ , glass wool filtration and swim-up (control). Total and motile sperm recovery rates were assessed, resuspended in fertilization TALP and incubated for 15-20 min in CO2 incubator. Samples prepared by each method were divided into two aliquots: one aliquot was studied for sperm quality (progressive motility, membrane integrity, viability, liveability), while the other was subjected to co-incubation with sets of 10-15 in vitro matured oocytes. Data on sperm quality were analysed by ANOVA, while in vitro fertilizing rates were compared by chi-squared test using SPSS-20. Least significant difference (LSD) test was used to compare treatment means. Glass wool filtration yielded higher total and motile sperm recovery rate, while Sephadex™ filtration improved (p < .05) sperm quality (progressive motility, membrane integrity, viability, liveability). Sperm preparation through Sephadex filtration yielded higher in vitro fertilization rate in terms of cleavage rate compared to glass wool filtration and swim-up (control). In conclusion, cryopreserved Nili-Ravi buffalo sperm selected through Sephadex filtration showed improved quality and yielded better fertilization rates (cleavage rate) of in vitro matured/fertilized oocytes. Sephadex filtration could be a promising technique for use in in vitro fertilization in buffalo.

Removal of hemangiosarcoma cells from canine blood with a cell salvage system and leukocyte reduction filter.

OBJECTIVE: To determine the ability of an intraoperative cell salvage (IOCS) system and a leukocyte reduction filter (LRF) to remove hemangiosarcoma (HSA) cells from canine blood. STUDY DESIGN: Cultured HSA cells were added to canine blood to simulate intraoperative hemorrhage and address hemoabdomen from ruptured splenic HSA. The blood/HSA cell mixture was processed through an IOCS, followed by LRF processing. SAMPLE POPULATION: Whole blood from 3 healthy dogs combined with cultured HSA cells. METHODS: The ability of quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), multiparameter flow cytometry, and cytologic examination to detect 50 HSA cells per milliliter of culture media was confirmed. RT-PCR, multiparameter flow cytometry, and cytologic examination were used to determine the presence of cultured HSA cells at 4 points during processing. RESULTS: HSA cells were found in all control samples and in all samples after IOCS but prior to LRF processing with all 3 cell detection methods. HSA cells were not found after IOCS/LRF processing with all 3 cell detection methods. CONCLUSION: IOCS combined with LRF processing is able to remove cultured HSA cells from canine blood. The addition of LRF to IOCS may allow application of IOCS in dogs with HSA. CLINICAL SIGNIFICANCE: A combination of IOCS and LRF processing may provide an alternative to allogeneic blood transfusion in dogs with hemoabdomen due to HSA.

A comparison of the effects of carbon dioxide and medical air for abdominal insufflation on respiratory parameters in xylazine-sedated sheep undergoing laparoscopic artificial insemination.

AIMS: To determine if abdominal insufflation with medical air will improve oxygenation and ventilation parameters when compared to insufflation with CO2 in xylazine-sedated sheep undergoing laparoscopic artificial insemination (AI). METHODS: Forty-seven sheep underwent oestrus synchronisation and were fasted for 24 hours prior to laparoscopic AI. Each animal was randomised to receive either CO2 or medical air for abdominal insufflation. An auricular arterial catheter was placed and utilised for serial blood sampling. Respiratory rates (RR) and arterial blood samples were collected at baseline, after xylazine (0.1 mg/kg I/V) sedation, 2 minutes after Trendelenburg positioning, 5 minutes after abdominal insufflation, and 10 minutes after being returned to a standing position. Blood samples were collected in heparinised syringes, stored on ice, and analysed for arterial pH, partial pressure of arterial O2 (PaO2), and CO2 (PaCO2). The number of ewes conceiving to AI was also determined. RESULTS: Repeated measures ANOVA demonstrated temporal effects on RR, PaO2, PaCO2 and arterial pH during the laparoscopic AI procedure (p<0.001), but no difference between insufflation groups (p>0.01). No sheep experienced hypercapnia (PaCO2>50 mmHg) or acidaemia (pH<7.35). Hypoxaemia (PaO2<70 mmHg) was diagnosed during the procedure in 14/22 (64%) ewes in the CO2 group compared with 8/23 (35%) ewes in the medical air group (p=0.053). Overall, 15/20 (75%) ewes in the CO2 group conceived to AI compared with 16/22 (72.7%) in the medical air group (p=0.867). CONCLUSIONS AND CLINICAL RELEVANCE: There were no statistical or clinical differences in RR, PaO2, PaCO2, pH, or conception to AI when comparing the effects of CO2 and medical air as abdominal insufflation gases. None of the sheep experienced hypercapnia or acidaemic, yet 42% (19/45) of sheep developed clinical hypoxaemia, with a higher percentage of ewes in the CO2 group developing hypoxaemia than in the medical air group. Based on the overall analysis, medical air could be utilised as a comparable alternative for abdominal insufflation during laparoscopic AI procedures.

Comparison of two methods of seminal plasma removal on buffalo (Bubalus bubalis) sperm cryopreservation.

Cryopreservation causes damage to spermatozoa, and methods minimizing this damage are therefore needed. Although much discussed, seminal plasma removal has become an alternative to improve sperm quality and viability after freezing and has been applied to different species in attempt to obtain good results. The objective of this study was to evaluate semen quality in buffaloes submitted to two methods for seminal plasma removal (filtration and centrifugation). Semen samples were collected from seven Murrah buffalo bulls (Bubalus bubalis) once a week for 8 weeks. Each ejaculate was divided into three groups: control (presence of seminal plasma), centrifugation and filtration. Sperm kinetics was evaluated with the computer-assisted sperm analysis (CASA) system. Plasmalemma and acrosomal membrane integrity, mitochondrial membrane potential and reactive oxygen species (ROS) were measured by flow cytometry, and lipid peroxidation was evaluated by the thiobarbituric acid reactive substances (TBARS) assay. Seminal plasma removal did not improve sperm kinetics compared to the control group. Centrifugation increased the number of cells with damaged acrosomal membranes (0.77 ± 0.05) and filtration caused greater plasmalemma and acrosomal membrane damage (22.18 ± 1.07). No difference in the mitochondrial membrane potential was observed between groups. In contrast, ROS production was higher in the centrifugation group compared to the control and filtration groups, although no differences in TBARS formation were detected. In conclusion, seminal plasma removal did not improve the quality of thawed buffalo semen compared to control in terms of sperm kinetics, membrane integrity, mitochondrial membrane potential or lipid peroxidation.

Short communication: Effects of nanofiltration and evaporation on the physiochemical properties of milk protein during processing of milk protein concentrate.

The aim of this work was to evaluate the effects of nanofiltration and evaporation concentration technologies on the physiochemical properties of milk protein concentrate (MPC) during processing. Skim milk, ultrafiltered milk, evaporated milk, nanofiltered milk, evaporated MPC, and nanofiltered MPC samples were collected at different processing stages. Chemical composition, microstructure of casein micelles, free sulfhydryl content, and surface hydrophobicity of the samples were determined. The insolubility index of MPC was also determined. Casein micelles aggregated compactly after evaporation while surface hydrophobicity increased and free sulfhydryl content decreased in evaporated milk compared with skim milk. However, the microstructure of the casein micelles was relatively undisturbed after nanofiltration, with reduced surface hydrophobicity and free sulfhydryl content. No significant difference was found in chemical composition between the 2 MPC preparations: approximately 61.40% protein and 28.49% lactose. In addition, the particulate microstructures of both MPC were similar. However, the insolubility index of evaporated MPC was significantly (0.58mL) higher than that of nanofiltered MPC. Nanofiltration may be an effective way to improve the solubility of MPC products.

Enumeration of clostridia in goat milk using an optimized membrane filtration technique.

A membrane filtration technique developed for counting butyric acid bacteria in cow milk was further developed for analysis of goat milk. Reduction of the sample volume, prolongation of incubation time after addition of proteolytic enzyme and detergent, and a novel step of ultrasonic treatment during incubation allowed filtration of goat milk even in the case of somatic cell counts (SCC) exceeding 10(6)/mL. However, filterability was impaired in milk from goats in late lactation. In total, spore counts were assessed in 329 farm bulk goat milk samples. Membrane filtration technique counts were lower than numbers revealed by the classic most probable number technique. Thus, method-specific thresholds for milk to evaluate the risk of late blowing have to be set. As expected, the spore counts of milk samples from suppliers not feeding silage were significantly lower than the spore counts of milk samples from suppliers using silage feeds. Not only were counts different, the clostridial spore population also varied significantly. By using 16S rRNA gene PCR and gene sequencing, 342 strains from 15 clostridial species were identified. The most common Clostridium species were Clostridium tyrobutyricum (40.4%), Clostridium sporogenes (38.3%), Clostridium bifermentans (7.6%), and Clostridium perfringens (5.3%). The 2 most frequently occurring species C. tyrobutyricum and C. sporogenes accounted for 84.7% of the isolates derived from samples of suppliers feeding silage (n=288). In contrast, in samples from suppliers without silage feeding (n=55), these species were detected in only 45.5% of the isolates.

An 18-µm microaggregate blood filter does not cause hemolysis during in vitro whole blood transfusions in sea turtles.

OBJECTIVE: Determine the hemolytic effect of an 18-µm microaggregate blood filter during in vitro sea turtle whole blood transfusions as well as describe the average diameter of leatherback (Dermochelys coriacea) and Kemp's ridley sea turtle (Lepidochelys kempii) RBCs. ANIMALS: 5 green (Chelonia mydas), 5 loggerhead (Caretta caretta), and 5 Kemp's ridley sea turtles (total n = 15). METHODS: Heparinized sea turtle blood was infused at 60 mL/h through a microbore extension set without and then with a postsyringe, inline 18-µm microaggregate blood filter. Pre- and postfiltration PCV, Hct, total solids, sodium, chloride, potassium, glucose, and free plasma hemoglobin concentrations were measured. With the use of light microscopy and archived blood smears, the maximum and minimum diameter of 20 RBCs from each of the 5 leatherback and 5 Kemp's ridley sea turtles were measured with a calibrated ocular micrometer using 400X magnification. RESULTS: There were no significant differences between pre- and postfiltration samples for Hct, total solids, sodium, chloride, potassium, glucose, and free plasma hemoglobin concentrations; however, there was a significant median postfiltration decrease in PCV of approximately 4%, representing a 13% decrease of the total RBCs transfused. Average maximum diameters for leatherback and Kemp's ridley sea turtle RBCs were 19.7 and 16.1 µm, respectively. CLINICAL RELEVANCE: Although the 18-µm microaggregate blood filter does not hemolyze transfused sea turtle RBCs and is likely safe for in vivo blood transfusions, the filter's pores may retain a small proportion of infused RBCs given their diameter.

Computational fluid dynamics of fish gill rakers during crossflow filtration.

We study crossflow filtration mechanisms in suspension-feeding fishes using computational fluid dynamics to model fluid flow and food particle movement in the vicinity of the gill rakers. During industrial and biological crossflow filtration, particles are retained when they remain suspended in the mainstream flow traveling across the filter surface rather than traveling perpendicularly to the filter. Here we identify physical parameters and hydrodynamic processes that determine food particle movement and retention inside the fish oral cavity. We demonstrate how five variables affect flow patterns and particle trajectories: (1) flow speed inside the fish oral cavity, (2) incident angle of the flow approaching the filter, (3) dimensions of filter structures, (4) particle size, and (5) particle density. Our study indicates that empirical experiments are needed to quantify flow parameters inside the oral cavity, and morphological research is needed to quantify dimensions of the filter apparatus such as gill rakers, the gaps between rakers, and downstream barriers. Ecological studies on suspension-feeding fishes are also needed to quantify food particle size and density, as these variables can affect particle retention due to hydrodynamic processes during crossflow filtration.

Short communication: Examination of milk filters by real-time PCR as a herd-level indicator of the presence of Mycobacterium avium subspecies paratuberculosis in dairy herds.

The aim of this study was to assess the suitability of real-time quantitative PCR (qPCR) for the detection of Mycobacterium avium ssp. paratuberculosis (MAP) in milk filters as a herd level indicator of paratuberculosis infection. Seventy-nine samples from textile or metal milk filters from 15 herds with defined MAP prevalence (infection status = noninfected, 0-5%, 5-10%, or >10% of animals with clinically confirmed paratuberculosis) were analyzed. The MAP DNA was isolated by a modified commercially available protocol for feces, and detection and quantification of the pathogen was performed by the IS900 qPCR. Mycobacterium avium ssp. paratuberculosis DNA was detected in 63 (79.7%) samples. Determination of MAP infection established by fecal and tissue culture was correctly confirmed by the analysis of milk filters on 11 of 12 infected farms; MAP was not detected in filters from 3 farms where paratuberculosis was never diagnosed. Statistical analysis of the data supports the evidence that milk filters can be used as a template for the direct detection of MAP on the herd level. The probability of successful MAP detection in milk filters in a herd with MAP-infected cows is at least 94.3%. Absolute numbers of MAP detected on the milk filter can be used for a rough estimation of paratuberculosis prevalence >10% in the herd. Analysis of milk filters for the presence of MAP can be a useful tool for the detection of paratuberculosis on the herd level before any individual control strategies.