Impact of corn silage or corn silage plus protein supplementation on the ingestive and rumination behaviours, ruminal fermentation characteristics and efficiency of grazing dairy cows.

This study's objective was to compare two options of pasture supplementation: corn silage (CS) alone or corn silage mixed with protein concentrate. The experiment was conducted with 18 lactating Holstein cows in mid-lactation in a crossover design that included three treatments and three data collection periods. All cows had access to pasture for 17 h/day with an average herbage allowance of 16 kg dry matter (DM)/cow/day and were offered in-barn corn silage, corn silage mixed with protein concentrate, or no supplementation. Cows were equipped with pH sensors residing in the reticulum and, during the 7-day data collection periods, with a jaw movement recorder. Nonsupplemented cows produced 21.3 kg energy-corrected milk (ECM) and ate 13.3 kg DM herbage at pasture. Cows supplemented with corn silage and corn silage plus protein produced 2.5 and 4.5 kg/day more ECM, respectively, consumed 3.4 and 3.3 kg/day more DM in total, respectively, ate for a shorter period of time, and ruminated longer than their nonsupplemented peers. Supplemented cows were almost able to cover their energy requirements and mobilised less body mass in contrast to the nonsupplemented cows. Cows offered corn silage plus protein showed increased ECM production, increased milk urea content and lower nitrogen use efficiency (NUE) compared to cows supplemented with corn silage only. Nonsupplemented dairy cows had the highest milk urea content and performed worst in terms of NUE. The best feed conversion efficiency resulted from the nonsupplemented dairy cows and those supplemented with corn silage plus protein. In nonsupplemented cows, the high feed conversion efficiency seemed to be due to the increased mobilisation of body mass. As a result of the starch-rich supplementations, the ruminal acetic:propionic acid ratio became smaller, and the proportions of n-butyric acid increased. The mean reticular pH values did not substantially vary across the three feeding treatments. For the choice of a supplementation option, herbage allowance and cost of supplement will have to be considered, but aspects of feed-food competition as well as animal welfare should not be ignored.

Influence of pre-grazing herbage mass on bite mass, eating behaviour, and dairy cow performance on pasture.

Knowledge about individual daily herbage dry matter (DM) intake (DMI) helps identifying efficient dairy cows and adapting supplementation better to herbage intake and nutrient requirements of grazing dairy cows. With the aid of behavioural characteristics, raw data recorded with the RumiWatch (RW) system and processed with the RW converter 0.7.3.31 (C31), estimation of herbage DMI may be possible. First, C31, which allows differentiation of prehension bites and mastication chews, was validated through direct observation of behavioural characteristics and compared to the previous RW converter 0.7.3.11 (C11). Further, the influence of a low and high pre-grazing herbage mass (HM), with the same target herbage allowance (HA), on bite mass, DMI, number of prehension bites, and milk production was investigated. In total, 24 lactating Holstein cows were pairwise allotted to one of two HM treatments. The cows received a new pasture paddock twice per day with a daily target HA of 22 kg DM per cow/day. On average, low HM (LHM) and high HM (HHM) paddocks had an HM of 589 and 2288 kg DM/ha, respectively, above 6.7 click units (1 CU = 0.5 cm). Overall, LHM cows produced 2.7 kg/day more milk and 2.5 kg/day more energy-corrected milk, had the same herbage DMI and a similar prehension bite mass. The averaged bite mass per week was 0.49 g DM/bite (LHM) or 0.47 g DM/bite (HHM), respectively. A longer eating time (617 vs. 559 min/day) and a shorter rumination time (297 vs. 365 min/day) were observed for the LHM cows compared with the HHM cows. The validation of the RW showed similar results for C11 and C31 apart from prehension bites, where C31 showed a mean absolute deviation of 12.4%. Pre-grazing HM had no effect on relevant behavioural characteristics for prospective intake estimation, namely, bite mass and number of prehension bites.

Holographic PIV/PTV for nano flow rates-A study in the 70 to 200 nL/min range.

Accurately measuring flow rates is a key requirement in many medical applications such as infusion and drug delivery systems. A major drawback of current systems is the low resolution of the sensors in the low flow rate regime. In this article, we present a method based on Holographic PIV/PTV that has been used for the first time to measure flow rates in the range of a few nL/min. Our method requires a very simple setup that combines lensless holography with particle velocimetry. For flow rates in the 70 to 200 nL/min range, the highest uncertainty was 5.6% (coverage factor k=2). This is an open-source project; the CAD designs and software source code are available at https://github.com/gui-miotto/holovel.

Magnetophoresis in Centrifugal Microfluidics at Continuous Rotation for Nucleic Acid Extraction.

Centrifugal microfluidics enables fully automated molecular diagnostics at the point-of-need. However, the integration of solid-phase nucleic acid extraction remains a challenge. Under this scope, we developed the magnetophoresis under continuous rotation for magnetic bead-based nucleic acid extraction. Four stationary permanent magnets are arranged above a cartridge, creating a magnetic field that enables the beads to be transported between the chambers of the extraction module under continuous rotation. The centrifugal force is maintained to avoid uncontrolled spreading of liquids. We concluded that below a frequency of 5 Hz, magnetic beads move radially inwards. In support of magnetophoresis, bead inertia and passive geometrical design features allow to control the azimuthal bead movement between chambers. We then demonstrated ferrimagnetic bead transfer in liquids with broad range of surface tension and density values. Furthermore, we extracted nucleic acids from lysed Anopheles gambiae mosquitoes reaching comparable results of eluate purity (LabDisk: A260/A280 = 1.6 ± 0.04; Reference: 1.8 ± 0.17), and RT-PCR of extracted RNA (LabDisk: Ct = 17.9 ± 1.6; Reference: Ct = 19.3 ± 1.7). Conclusively, magnetophoresis at continuous rotation enables easy cartridge integration and nucleic acid extraction at the point-of-need with high yield and purity.

OralDisk: A Chair-Side Compatible Molecular Platform Using Whole Saliva for Monitoring Oral Health at the Dental Practice.

Periodontitis and dental caries are two major bacterially induced, non-communicable diseases that cause the deterioration of oral health, with implications in patients' general health. Early, precise diagnosis and personalized monitoring are essential for the efficient prevention and management of these diseases. Here, we present a disk-shaped microfluidic platform (OralDisk) compatible with chair-side use that enables analysis of non-invasively collected whole saliva samples and molecular-based detection of ten bacteria: seven periodontitis-associated (Aggregatibacter actinomycetemcomitans, Campylobacter rectus, Fusobacterium nucleatum, Prevotella intermedia, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola) and three caries-associated (oral Lactobacilli, Streptococcus mutans, Streptococcus sobrinus). Each OralDisk test required 400 µL of homogenized whole saliva. The automated workflow included bacterial DNA extraction, purification and hydrolysis probe real-time PCR detection of the target pathogens. All reagents were pre-stored within the disk and sample-to-answer processing took < 3 h using a compact, customized processing device. A technical feasibility study (25 OralDisks) was conducted using samples from healthy, periodontitis and caries patients. The comparison of the OralDisk with a lab-based reference method revealed a ~90% agreement amongst targets detected as positive and negative. This shows the OralDisk's potential and suitability for inclusion in larger prospective implementation studies in dental care settings.

RespiDisk: a point-of-care platform for fully automated detection of respiratory tract infection pathogens in clinical samples.

We present the RespiDisk enabling the fully automated and multiplex point-of-care (POC) detection of (currently) up to 19 respiratory tract infection (RTI) pathogens from a single sample based on reverse transcriptase polymerase chain reaction (RT-PCR). RespiDisk comprises a RTI-specific implementation of the centrifugal microfluidic LabDisk platform and combines new and existing advanced unit operations for liquid control, thereby automating all assay steps only by a spinning frequency and temperature protocol in combination with the use of a permanent magnet for in situ bead handing. The capabilities of the system were demonstrated with 36 tested quality samples mimicking clinical conditions (clinical and/or cultured material suspended in transport medium or synthetic bronchoalveolar lavage (BAL)) from past external quality assessment (EQA) panels covering 13 of the 19 integrated RTI detection assays. In total, 36 samples × 19 assays/sample resulting in 684 assays were performed with the RespiDisk, and its analytical performance was in full agreement with the routine clinical workflow serving as reference. A strong feature of the platform is its universality since its components allow the simultaneous detection of a broad panel of bacteria and viruses in a single run, thereby enabling the differentiation between antibiotic-treatable diseases. Furthermore, the full integration of all necessary biochemical components enables a reduction of the hands-on time from manual to automated sample-to-answer analysis to about 5 min. The study was performed on an air-heated LabDisk Player instrument with a time-to-result of 200 min.

Real-time stability testing of air-dried primers and fluorogenic hydrolysis probes stabilized by trehalose and xanthan.

A method for conserving primers and differently labeled fluorogenic hydrolysis (i.e., TaqMan) probes at ambient conditions is presented. Primers and hydrolysis probes with four different fluorophore-quencher combinations (6- FAM-BHQ1, HEX-BHQ1, ROX-BHQ650, and Cy5-BHQ2) were mixed with trehalose and xanthan at final concentrations of 56 mM and 2.78 mM, respectively. Mixtures were air-dried at 23°C for 30 min on strips composed of cyclo olefin polymer (COP), a material widely used in the manufacturing of in vitro diagnostic (IVD) test carriers. After one year of storage, the functionality of the primers and fluorophore-quencher combinations was validated by real-time polymerase chain reaction (real-time PCR), confirming their stability when stored in the presence of stabilizers, with the best results achieved using trehalose. This approach could be of great benefit for manufacturing IVD systems, for example, for genotyping applications based on multiplexing using different fluorescent dyes.