Milk Samples Research Articles

The concentration of maternal sacubitril/valsartan transferred into human milk is negligible.

Peripartum cardiomyopathy (PPCM) is a common cause of heart failure (HF) in the peripartum. Some medications are considered safe while breastfeeding. However, sacubitril/valsartan (Entresto), while efficacious, is not recommended in breastfeeding women due to concerns about adverse infant development, and no published data suggest otherwise. This study aimed to assess the transfer of sacubitril/valsartan into human milk and evaluate the infant's risk of drug exposure. The InfantRisk Human Milk Biorepository released samples and corresponding health information from five breastfeeding maternal-infant dyads exposed to sacubitril/valsartan. Sacubitril, valsartan, and LBQ657 (sacubitril active metabolite) concentrations were determined using liquid chromatography-mass spectrometry (LC/MS/MS) from timed samples 0, 1, 2, 4, 6, 8, 10, and 12 h following medication administration at steady state conditions. Valsartan levels were below the detection limit of 0.19 ng/mL in all milk samples. Sacubitril was measurable in all milk samples of the five participants, peaking 1 h after drug administration at a mean concentration of 1.52 ng/mL for a total infant dose of 0.00049 mg/kg/12 h and a relative infant dose (RID) calculated at 0.01%. The maximum concentration of its active metabolite LBQ657 in the milk samples was observed 4 h after medication administration and declined over the remaining 12-h dosing interval, for an average concentration of 9.5 ng/mL. The total infant dose was 0.00071 mg/kg/12 h, and the RID was 0.22%. Two mothers reported continuing to breastfeed while taking sacubitril/valsartan; both mothers stated observing no negative effects in their breastfed infants. The transfer of sacubitril/valsartan into human milk is minimal. These concentrations are unlikely to pose a significant risk to breastfeeding infants, with a combined calculated RID of <0.25%, which is far lower than the industry safety standards (RID <10%).

Insights into the early-life chemical exposome of Nigerian infants and potential correlations with the developing gut microbiome

Early-life exposure to natural and synthetic chemicals can impact acute and chronic health conditions. Here, a suspect screening workflow anchored on high-resolution mass spectrometry was applied to elucidate xenobiotics in breast milk and matching stool samples collected from Nigerian mother-infant pairs (n = 11) at three time points. Potential correlations between xenobiotic exposure and the developing gut microbiome, as determined by 16S rRNA gene amplicon sequencing, were subsequently explored. Overall, 12,192 and 16,461 features were acquired in the breast milk and stool samples, respectively. Following quality control and suspect screening, 562 and 864 features remained, respectively, with 149 of these features present in both matrices. Taking advantage of 242 authentic reference standards measured for confirmatory purposes of food bio-actives and toxicants, 34 features in breast milk and 68 features in stool were identified and semi-quantified. Moreover, 51 and 78 features were annotated with spectral library matching, as well as 416 and 652 by in silico fragmentation tools in breast milk and stool, respectively. The analytical workflow proved its versatility to simultaneously determine a diverse panel of chemical classes including mycotoxins, endocrine-disrupting chemicals (EDCs), antibiotics, plasticizers, perfluorinated alkylated substances (PFAS), and pesticides, although it was originally optimized for polyphenols. Spearman rank correlation of the identified features revealed significant correlations between chemicals of the same classification such as polyphenols. One-way ANOVA and differential abundance analysis of the data obtained from stool samples revealed that molecules of plant-based origin elevated as complementary foods were introduced to the infants’ diets. Annotated compounds in the stool, such as tricetin, positively correlated with the genus Blautia. Moreover, vulgaxanthin negatively correlated with Escherichia-Shigella. Despite the limited sample size, this exploratory study provides high-quality exposure data of matched biospecimens obtained from mother-infant pairs in sub-Saharan Africa and shows potential correlations between the chemical exposome and the gut microbiome.

Application of an electronic nose for the diagnosis of ketosis in dairy cows

Ketosis in dairy cows is a nutritional and metabolic disease that severely affects the milk industry and dairy cow health. An electronic nose can identify volatile organic compounds (VOCs). This study applied the electronic nose to detect VOCs in dairy cow milk, feces, and blood. Blood ketone bodies (BHBA) were measured, and 10 ketosis cows, 71 subclinical ketosis cows, and 75 healthy cows were considered the research subjects. Cow milk, feces, and blood samples were collected for electronic nose testing. The measured VOC data were processed through multivariate analysis and machine learning algorithms. The electronic nose analysis results were validated through the GC-MS analysis of the milk samples. The results unveiled spatial differences in the electronic nose VOC values of milk, feces, and blood, and the cumulative contribution rates of both principal component variables (PC) 1 and PC2 reached more than 95%. The machine learning algorithm results revealed that the accuracy of the milk probabilistic neural network (PNN) was 87.5%, the stool support vector machine was 100%, and the blood PNN was 66.7%. This result revealed that electronic nose detection can effectively differentiate healthy, subclinical ketosis, and clinical ketosis cows. According to the milk GC-MS results, significant differences were noted in the VOC content of milk samples between the different groups.The aforementioned studies revealed that electronic nose can aid in the effective prediction and diagnosis of ketosis in dairy cows. This lays the foundation for establishing a new method for diagnosing ketosis in dairy cows.

Subclinical mastitis in Jersey dairy cows and its effects on productivity and inflammatory markers

This study assessed the effect of subclinical mastitis on the productivity and inflammatory markers in Jersey dairy cows. Blood, milk, and milk yield data were collected from 59 Jersey dairy cows reared under a semi-extensive system. Milk samples were collected from individual collectors and evaluated for their somatic cell count (SCC), lactose (Lact), protein, fat, total and defatted dry extracts (DDE), casein, freezing point, and milk urea nitrogen (MUN) levels. After milking, blood was collected by puncturing the coccygeal arteriovenous complex. In the serum samples, the inflammatory biomarkers paraoxonase-1, albumin, and total plasma protein levels were analyzed using colorimetric methods. Samples of the entire diet provided and pastures were submitted for bromatological analysis. Additionally, the body condition score, number of lactations, milk yield, days of lactation, calving interval, number of inseminations until conception, calving-conception interval, and days of pregnancy were analyzed. The cows were categorized based on their SCC into a subclinical mastitis group (SubG), with SCC levels greater than or equal to 200,000 cells/mL, and a control group (CG), with SCC levels below 200,000 cells/mL. Blood metabolic variables, milk quality, and milk production data were analyzed using multivariate regression, analysis of variance, and logistic regression using the SAS® program. The multivariate regression analysis revealed that several markers, such as Lact, total protein, and casein, influenced milk composition, predicting over 97% of the data. The SubG showed lower concentrations of Lact (4.37 vs. 4.47%; P = 0.0002), MUN (20.55 vs. 23.85 mg/dL; P = 0.02), and DDE (9.24 vs. 9.50%; P = 0.02) compared with the CG; moreover, animals in the SubG had a higher number of lactations compared with those in the CG (4.30 vs. 2.69; P = 0.0039). Cows with a Lact content lower than 4.265% (quartile 25%), compared with those with Lact content greater than 4.565% (quartile 100%), were 52 times more likely to have subclinical mastitis. Jersey dairy cows with subclinical mastitis have lower Lact, urea, and defatted dry extract levels in their milk as well as a higher number of lactations. Therefore, Lact levels in milk serve as a good diagnostic marker of subclinical mastitis in Jersey cows. Subclinical mastitis in Jersey dairy cows did not decrease milk yield nor effect reproductive performance.

Non-targeted mass spectrometry and feature-based molecular networking for determination of branched fatty acid esters of hydroxy fatty acids in milk.

Branched fatty acid esters of hydroxy fatty acids (FAHFAs) represent trace lipids with significant natural biological functions. While exogenous FAHFAs have been extensively studied, research on FAHFAs in milk remains limited, constraining our grasp of their nutritional roles. This study introduces a non-targeted mass spectrometry approach combined with chemical networking of spectral fragmentation patterns to uncover FAHFAs. Through meticulous sample handling and comparisons of various data acquisition and processing modes, we validate the method's superiority, identifying twice as many FAHFAs compared to alternative techniques. This validated method was then applied to different milk samples, revealing 45 chemical signals associated with known and potential FAHFAs, alongside findings of 66 ceramide/hexosylceramide (Cer/HexCer), 48 phosphatidyl ethanolamine/lyso phosphatidyl ethanolamine (PE/LPE), 21 phosphatidylcholine/lysophosphatidylcholine (PC/LPC), 16 phosphatidylinositol (PI), 7 phosphatidylserine (PS), and 11 sphingomyelin (SM) compounds. This study expands our understanding of the FAHFA family in milk and provides a fast and convenient method for identifying FAHFAs.

MEASUREMENT OF MICROBIOLOGICAL QUALITY OF RAW GOAT´S MILK BY LASER FLOW CYTOMETRY

The popularity of goat's milk and its products is growing in Slovakia. Among the mandatory health safety features of raw goat's milk is the total bacterial count. It can be determined by the anchor plate-cultivation method, which is, however, tedious, and laborious. Laser flow cytometry is a modern alternative method for measuring the microbiological quality of raw milk. To use the results from laser flow cytometry method for legislative purposes, it is necessary to create a representative conversion of the primarily measured results obtained by this method into the CFU/mL scale. This work describes the creation of such a conversion for raw goat's milk, which will be valid for the Slovak region. After measuring several hundred individual and bulk tank samples of raw goat's milk on a BactoScan FC laser flow cytometer and at the same time using the anchor method according to STN EN ISO 4833-1 and after their subsequent logarithmic transformation, such a conversion was created in the form of log10(CFU/mL) = 1.0174 x log10(IBC/ml) + 2.7483. Subsequently, this was verified by comparing the recalculated and directly measured results in CFU/mL, using additional data set of samples. The influence of the somatic cell count in raw goat's milk, as a possible interfering factor, was also assessed.

Commensal bacteria in raw milk, the reservoirs for antibiotic resistance genes during cold chain transportation

The emergence and prevalence of antibiotic resistance genes (ARGs) in food pose a great threat to public health and have attracted globally attention. Shotgun metagenomics sequencing is widely used to characterize ARGs in the environment. However, the distribution, co-occurrence patterns, and host information of ARGs in raw milk to the cold chain transportation duration with shotgun metagenomics sequencing, are not fully understood in China. In this study, species annotation and ARG annotation were performed on 40 raw milk samples collected from Beijing (BJ), Hebei (HB), Inner Mongolia (NMG), Shanghai (SH), and Guangdong (GD) using shotgun sequencing based on the Illumina platform. A total of 4731 bacterial species were annotated, of which 37 were dominant (relative abundance >1%). And also, 259 ARGs were annotated, corresponding to 69 kinds of antibiotic resistance. The NMG samples, which had the lowest bacterial community richness and bacterial diversity, were annotated with the highest number of ARGs, revealing the potential risk of bacterial resistance in NMG samples. Spearman correlation analysis and network analysis were showed that the probable bacterial hosts of multiple highly abundant ARGs (MacB and VanRC) in samples from provinces except for the BJ samples were Pseudomonas and Serratia, rather than conventionally known foodborne pathogens. It is noteworthy that the relative abundance of Pseudomonas fluorescens conformed to the Allometricl function with the cold chain transportation duration, the finding that could provide a reference for assessing the freshness and the risk assessment of antibiotic resistance genes of raw milk. In addition, the dairy industry should be aware of the risk of ARGs transferring from highly abundant commensal bacteria. Raw milk for transportation by cold chain, higher abundance of Pseudomonas in some samples should be kept in mind, and the roles of ARGs carried by these bacteria should be further investigated.

Persistent Organic Contaminants in Human Milk

Persistent organic pollutants (POPs) are always components of human milk and can be transferred to infants through lactation. This paper describes the POPs and discusses the risks that POPs in human milk pose to nursing infants. FeaturesofselectPOPsthatareexplainedinthispaperincludetheiruses,toxicityandtemporaltrends. The levels of most POPs are declining in human milk due to a series of international bans and restrictions that have been placed on the manufacturing and/or use of the chemicals. However, there is still concern regarding the continued exposure to POPs from leaks in electrical equipment and vector control programs. Although these contaminants continue to be detected in human milk samples, women are still encouraged to breastfeed their newborn children. It is firmly believed that the benefits of lactation for both the mother and her child outweigh the health hazards that POPs present.

Assessment of Heavy Metals and Trace Elements in the Human Milk of Women Living in Latvia and an Evaluation of Influencing Factors.

During lactation, heavy metals and trace elements can be mobilised from the maternal body stores and excreted via human milk. A total of 66 mature human milk samples were collected from lactating women in Latvia between 2016 and 2017 to analyse the content of As, Cd, Pb, Al, Sn, and Ni. Additionally, 50 mature human milk samples were collected between 2022 and 2023 to analyse the content of Cd and Pb. The content of heavy metals and trace elements in human milk was determined using ICP-MS. Only two individual human milk samples contained heavy metals above the method's detection limit-one with an arsenic content of 0.009 mg kg-1 and one with a lead content of 0.047 mg kg-1. The preliminary data show that human milk among lactating women in Latvia contains only insignificant amounts of heavy metals and trace elements. Concern over such content should not be a reason to choose formula feeding over breastfeeding. Nevertheless, heavy metals, trace elements and other pollutants in human milk should be continuously monitored.

Transformation of heavy metals from contaminated water to soil, fodder and animals

A serious environmental problem that threatens soil quality, agricultural productivity, and food safety is heavy metal pollution in water sources. Heavy metal pollution is the main problem in tehsil Pasrur, Sialkot, Pakistan. Present study was arranged to notice the heavy metals in water, soil, forages and buffalo milk. There are seven sites that were used for this experiment. Highest malondialdehyde (MDA) contents (3.00 ± 0.01) were noticed in barseem roots at site 7. Ascorbate Peroxidase (APX) was reached at its peak (1.93 ± 0.01) at site 7 in the fresh barseem. Maximum protein contents (0.36 ± 0.01) were observed in fresh plant samples at site 2. Site 3's buffalo milk samples had the highest Ni content (7.22 ± 0.33 ppm), while Site 3's soil samples had the lowest Cr content (8.89 ± 0.56 ppm), Site 1's plant shoots had the lowest Cr content (27.75 ± 1.98 ppm), and Site 3's water had the highest Cr content (40.07 ± 0.49 ppm). The maximum fat content (5.38 ± 2.32%) was found in the milk of the animals at site 7. The highest density (31.88 ± 6.501%), protein content (3.64 ± 0.33%), lactose content (5.54 ± 0.320%), salt content (0.66 ± 0.1673%), and freezing point (− 0.5814 ± 0.1827 °C) were also observed in the milk from animals at site 7, whereas site 5 displayed the highest water content (0.66 ± 0.1673%) and peak pH value (11.64 ± 0.09). In selected samples, the pollution load index for Ni (which ranged from 0.01 to 1.03 mg/kg) was greater than 1. Site 7 has the highest conductivity value (5.48 ± 0.48). Values for the health risk index varied from 0.000151 to 1.00010 mg/kg, suggesting that eating tainted animal feed may pose health concerns. Significant health concerns arise from metal deposition in the food chain from soil to feed, with nickel having the highest health risk index.

Organophosphate esters in milk across thirteen countries from 2020 to 2023: Concentrations, sources, temporal trends and ToxPi priority to humans

Recent increases in organophosphate ester (OPE) application have led to their widespread presence, yet little is known about their temporal trends in food. This study collected milk samples from 13 countries across three continents during 2020–2023, finding detectable OPEs in all samples (range: 2.25–19.7; median: 7.06 ng/g ww). Although no statistical temporal differences were found for the total OPEs during the 4-year sampling campaign, it was interesting to observe significant variations in the decreasing trend for Cl-OPEs and concentration variations for aryl-OPEs and alkyl-OPEs (p < 0.05), indicating changing OPE use patterns. Packaged milk exhibited significant higher OPE levels than those found in directly collected raw unpackaged milk, and milk with longer shelf-life showed higher OPE levels, revealing packaging material as a contamination source. No significant geographical differences were observed in milk across countries (p > 0.05), but Shandong Province, a major OPE production site in China, showed relatively higher OPE concentrations. The Monte Carlo simulation of estimated daily intakes indicated no exposure risk from OPEs through milk consumption. The molecular docking method was used to assess human hormone binding affinity with OPEs, amongst which aryl-OPEs had the highest binding energies. The Toxicological-Priority-Index method which integrated chemical property, detection frequency, risk quotients, hazardous quotients and endocrine-disrupting effects was employed to prioritize OPEs. Aryl-OPEs showed the highest scores, deserving attention in the future.

Effects of Brown Seaweed (Ascophyllum nodosum) Supplementation on Enteric Methane Emissions, Metabolic Status and Milk Composition in Peak-Lactating Holstein Cows.

The dairy industry contributes significantly to anthropogenic methane emissions, which have an impact on global warming. This study aimed to investigate the effects of a dietary inclusion of brown seaweed Ascophyllum nodosum on enteric methane emissions (EMEs), hematological and blood biochemical profiles, and milk composition in dairy cows. Eighteen Holstein cows were divided into three groups: CON (non-supplemented cows), BS50 (50 mL of 10% A. nodosum), and BS100 (100 mL of 10% A. nodosum). In each cow, measurements of EME, dry matter intake (DMI), and milk yield (MY), as well as blood and milk sampling with respective analyzes, were performed before supplementation (P1), after 15 (P2) days, and after 30 (P3) days of supplementation. A. nodosum reduced (p < 0.05) methane production, methane yield, and methane intensity in both BS50 and BS100, and raised DMI (p < 0.05) only in BS50. Total bilirubin (p < 0.05) was higher in BS50 compared to CON cows in P2, and triacylglycerols were lower (p < 0.05) in BS50 than in CON cows in P3. Higher milk fat content was found in BS50 than in CON cows in P3. C16:0 proportions were higher (p < 0.05) in BS50 and BS100 than in CON cows, while C18:3n-3 was higher (p < 0.05) in BS100 than in BS50 and CON cows in P3. Dietary treatment with A. nodosum reduced EMEs and showed the potential to increase DMI and to improve energy status as well as milk composition in peak-lactating dairy cows.

Seasonal Variation and Dietary Effects in the Fatty Acid Composition of Bulk Tank Milk in Taiwan

Background: Information about seasonal variation in the fatty acid (FA) composition of bulk tak milk and its relationship with dietary factors in Taiwan remains to be completed. This study aimed to determine the average FA composition of milk in different seasons and collect dietary information to investigate the relationship between dietary characteristics and lactation performance, including FA concentration in bulk tank milk. Methods: Bulk tank milk samples were collected monthly from 90 dairy farms across Taiwan for two years to capture seasonal variations. Additionally, 36 bulk tank milk samples from three farms were collected monthly from August 2022 to July 2023. Result: Milk production (kg/day), fat (%), crude protein (%), true protein (%), lactose (%) and solids-not-fat (%) were significantly higher in the cold season than in the hot and warm seasons (P less than 0.05) over the studied period. In contrast, somatic cell counts were significantly lower in the cold season than in the hot and warm seasons (P less than 0.05). Total saturated FA and C14:0 concentrations were significantly higher in the cold season than in the hot and warm seasons (P less than 0.05). In contrast, the C18:1 concentration was significantly higher in the hot and warm seasons than in the cold season (P less than 0.05). However, the C18:0 concentration did not differ significantly by season. Fat and FA concentrations varied significantly among three selected dairy farms (P less than 0.05). Milk fat only differed significantly between the three farms in the hot season, with Farm B having the lowest milk fat concentration (P less than 0.05). FA concentrations showed significant seasonal effects at all three farms. These results provide an overview of milk FA variation and identify the critical factors influencing variation in bulk tank milk fat and individual farm FA concentrations: dietary ether extract content and forage choice.

Electrochemical detection platform based on graphene-zinc oxide composite nanomaterials for 17β-estradiol determination in milk

This research presents the development of an innovative electrochemical detection platform using a graphene-zinc oxide (G-ZnO) nanocomposite for the quantification of 17β-estradiol in milk. The G-ZnO nanocomposite, synthesized via a facile one-pot method, exhibited a unique wrinkled structure with a high specific surface area of 143 m²/g. Characterization through SEM, TEM, and XRD confirmed the effective integration of ultrafine ZnO nanoparticles onto graphene sheets. The electrochemical performance of the G-ZnO modified glassy carbon electrode was evaluated, revealing a notably low detection limit of 8.3 nM for 17β-estradiol, a significant improvement over traditional sensors. The sensor demonstrated a wide linear range of 0.05–100 μM and exceptional reproducibility with less than 3.5 % RSD over repeated measurements. Recovery studies in spiked milk samples yielded highly accurate results, with recovery percentages ranging from 97.8 % to 104.6 %. These findings underscore the potential of G-ZnO nanocomposites in creating sensitive, efficient, and reliable sensors for healthcare and environmental applications.

Quantitation of milk proteins in thermally treated milk samples and commercial food products by ELISA test kits

This study evaluated four ELISA kits for quantitation of milk proteins in thermally treated milk samples and food products. How reference materials may be used for comparison of kit performance was examined. Protein contents determined by Veratox Total Milk generally reflected those determined by the 660 nm total protein assay. BioKits BLG Kit was less affected by thermal treatment but resulted in overestimation of protein contents in samples that were boiled, autoclaved or dry-heated at ≤149 °C, while ELISA Systems Casein (ES Casein) and Beta-Lactoglobulin (ES BLG) assays underestimated protein levels in these samples. The four kits gave similar results for ice cream. Veratox registered higher concentrations in all products tested but its sensitivity was greatly lowered in retorted products. ES Casein underperformed Veratox for baked and retorted products. BioKits BLG maintained a better sensitivity towards fried, baked and retorted products while ES BLG exhibited reduced sensitivity for these products.

Ultrasensitive detection of Salmonella typhi using a PAM-free Cas14a-based biosensor

The effectiveness of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas14a1, widely utilized for pathogenic microorganism detection, has been limited by the requirement of a protospacer adjacent motif (PAM) on the target DNA strands. To overcome this limitation, this study developed a Single Primer isothermal amplification integrated-Cas14a1 biosensor (SPCas) for detecting Salmonella typhi that does not rely on a PAM sequence. The SPCas biosensor utilizes a novel primer design featuring an RNA-DNA primer and a 3′-biotin-modified primer capable of binding to the same single-stranded DNA (ssDNA) in the presence of the target gene. The RNA-DNA primer undergoes amplification and is blocked at the biotin-modified end. Subsequently, strand replacement is initiated to generate ssDNA assisted by RNase H and Bst enzymes, which activate the trans-cleavage activity of Cas14a1 even in the absence of a PAM sequence. Leveraging both cyclic chain replacement reaction amplification and Cas14a1 trans-cleavage activity, the SPCas biosensor exhibits a remarkable diagnostic sensitivity of 5 CFU/mL. Additionally, in the assessment of 20 milk samples, the SPCas platform demonstrated 100% diagnostic accuracy, which is consistent with the gold standard qPCR. This platform introduces a novel approach for developing innovative CRISPR-Cas-dependent biosensors without a PAM sequence.

The Influence of Milk Leakage, Udder Pressure and Further Risk Factors on the Development of New Intramammary Infections during the Dry Period of Dairy Cows.

Prevention of new intramammary infection (NIMI) during the dry period (DP) is essential to prevent the development of mastitis in dairy cows. To investigate risk factors for NIMI, 212 cows, comprising a total of 848 udder quarters, were examined in this study. Quarter milk samples were taken on the day of drying off and 7 ± 3 days after calving. Cow- and quarter-level associated risk factors were assessed at the beginning of the DP and after calving. In total, 7.1% of the udder quarters developed an NIMI between the samplings. Non-aureus staphylococci (40.4%) and Gram-negative pathogens (22.8%) were most frequently the cause of NIMI. The observed milk leakage prevalence was 16.7%, with a peak 24 h after drying off. Simultaneously, the udder pressure peaked 24 h after drying off. A significant correlation between milk yield on the day before drying off and milk leakage could be proven. Cows with quarters leaking milk produced an average milk yield of 28.32 kg on the day before drying off. Generalised linear mixed models and odds ratios were calculated to determine the significant risk factors for NIMI during the DP and early lactation. Quarters leaking milk had 3.4 higher odds for NIMI between the samplings compared to quarters without milk leakage. Quarters from cows with dirty udders had 3.1 higher odds of developing an NIMI between the samplings compared to quarters from cows with clean udders. The results of this study demonstrate the importance of dry cow management before drying off and during the critical period of active involution of the udder tissue.

An exonuclease I-assisted quencher-free 2-aminopurine aptasensor based on a multipath paper-based device for ultrasensitive detection of kanamycin

A quencher-free multipath microfluidic paper-based analytical device (µPAD) was constructed for ultrasensitive detection of kanamycin based on exonuclease I (Exo I)-assisted signal enhancement of 2-aminopurine (2-AP). Here, Exo I, a single-stranded DNA-specific nuclease, was introduced to fully liberate 2-AP mononucleotides to greatly enhance biosensing sensitivity. 2-AP, a fluorescent adenine analogue embedded in single-stranded DNA (ssDNA), was employed as the detection signal source. The fluorescence of 2-AP is strong in the mononucleotide state, while it can have low fluorescence and even no fluorescence in ssDNA and dsDNA, respectively. The 2-AP fluorescence probe included 2-AP DNA and kanamycin aptamer. When kanamycin was present, binding occurred between kanamycin and the aptamer, leading to ssDNA, which was further digested by Exo I. In this case, free 2-AP mononucleotides were liberated, indicating strong fluorescence. In addition, the captured kanamycin was released for binding with the new aptamer, which resulted in the formation of a binding-hydrolysis-release cycle with the aid of Exo I. Under optimal conditions, this µPAD exhibited sensitive and multipath detection of kanamycin at concentrations as low as 1.26 × 10−14 M with a wide range of 10−13–10−7 M. Furthermore, satisfactory results were achieved for analysing spiked kanamycin in milk and honey samples. This strategy is a very promising tool for monitoring antibiotics and evaluating the safety of animal-derived foods.

Efficacy of Green Silver Nanoparticles (GAbNPs) against Multidrug Resistance in Staphylococcus aureus from Mastitis-infected Cows

Background: Antimicrobial resistance being the major emerging problem for management of mastitis in cow hence, the research work was undertaken in a geographical area around Shirwal town in Satara district of Western Maharashtra, India to investigate the occurrence of multidrug resistance in Staphylococcus aureus (MDRSA) isolated from mastitic cow milk samples. Further assessment was carried out for antimicrobial efficacy of formulation of Green Silver Nanoparticles (GAgNPs) synthesized from Azadirechta indica against MDRSA of mastitis origin. Methods: A total of 250 milk samples from cows suspected of mastitis were screened by the California Mastitis Test (CMT). The CMT-positive samples were subjected to bacteriological investigation for the presence of S. aureus as an etiological agent. The isolates identified as S. aureus were further authenticated by the molecular method targeting 16s-rRNA and nuc gene. All isolates of S. aureus were subjected to an antimicrobial susceptibility test. The randomly selected MDRSA isolates were subjected to antimicrobial susceptibility testing with GAgNPs formulation were prepared from silver nitrate by reducing with Azadirachta indica leaf extract. Result: Out of 250 cow milk samples suspected for having mastitis were screened with CMT, 170 tested positive. From these milk samples 145 isolates of S. aureus were isolated and bacteriologically identified. When processed for antibiotic sensitivity 60 were detected having multidrug resistance. Maximum resistance was shown against beta-lactam antibiotics. GAgNP synthesized and characterized with visual appearance, spectroscopy and TEM (Transmission electron microscopy) and tested for in vitro antimicrobial efficacy against MDRSA showed significant antimicrobial property.

Exogenous xylanase increases total tract digestibility of energy and fiber in diets for gestating and lactating sows, but does not influence reproductive performance of sows

The hypothesis that exogenous xylanase added to diets for gestating and lactating sows will increase the apparent total tract digestibility (ATTD) of gross energy (GE) and total dietary fiber (TDF), increase digestible energy (DE) and metabolizable energy, and improve the reproductive performance of sows was tested. Two diets for gestating sows and two diets for lactating sows were formulated without or with 100 g per ton of an exogenous xylanase (16,000 units/kg). Diets were fed in two consecutive reproductive cycles. From 106 animals, 48 pregnant sows, organized in 4 blocks of 12 sows (6 sows per treatment in each block), were placed for 10 days in metabolism crates starting on day 35 (mid-gestation) and again on day 95 (late-gestation) with feces and urine being collected for 4 days. Sows were moved to the lactation unit on day 106 of gestation and feeding of lactation diets was initiated. Fecal samples were collected (grab-sampling) from days 10–14 post-farrowing. The number and weight of pigs born, mummified, stillborn, and weaned per sow were recorded, and survival rate and litter average daily gain were calculated. Litters were weaned on day 20 ± 1. All animals were rebred and 46 sows were placed in metabolism crates in mid and late-gestation as in the first cycle, and treatments in the farrowing unit during the second cycle were also as in the first cycle; however, colostrum and milk samples were collected from sows in the second cycle. Results indicated that reproductive performance was not different between sows fed control diets and sows fed diets with xylanase during the two reproductive cycles. In the first gestation period, the ATTD of TDF in late-gestation was greater (P < 0.05) in sows fed the diet with xylanase than in sows fed the control diet. During the first lactation, sows fed the diet with xylanase had greater (P < 0.05) ATTD of GE and TDF, and greater (P < 0.05) DE than sows fed the control diet. During the second gestation, sows fed the diet with xylanase had greater (P < 0.05) DE in mid-gestation. During the second lactation, sows fed the diet with xylanase had greater (P < 0.05) ATTD of GE and TDF, and greater (P < 0.05) DE than sows fed the control diet. In conclusion, DE was greater in diets with xylanase than in control diets during the two reproductive cycles, and sows fed the lactation diet with xylanase had greater digestibility of fiber than sows fed the control diet.