Cow Milk Samples Research Articles

Seasonal effects on urinary iodine concentrations in women of reproductive age: An observational study in Tanzania and South Africa

Iodine intake in populations is usually assessed by measuring urinary iodine concentrations (UICs) in spot samples. Hot climate conditions may reduce urine volume, thus leading to overestimations of UIC and thereby masking inadequate iodine intake. We investigated the effects of season on UICs in 2 populations exposed to high-temperature climates. In this observational study, we examined women (18-49 years) in Tanzania (ncold=206; nhot=179) and South Africa (ncold=157; nhot=126) during cold and hot seasons. From each woman in both seasons, we obtained two 24-hour urine collections and 2 spot urine samples, as well as salt, water, and cow's milk samples. We measured the urine volume, UIC, and urinary creatinine concentration (UCC). The 24-hour urinary iodine excretion (UIE) was calculated and used to estimate the iodine intake. We used linear mixed-effects models to test for differences between seasons. In Tanzanian women, we observed no seasonal effect on the urine volume, 24-hour UIE, 24-hour UIC, spot UIC, spot UIC:UCC ratio, or salt iodine concentration. In South African women, the median 24-hour urine volume was 1.40 L (IQR, 0.96-2.05 L) in the winter and 15% lower in the summer (P<0.001). The median 24-hour UIE was 184 µg/day (IQR, 109-267µg/day) in the winter and 34% lower in the summer (P<0.001), indicating a lower iodine intake. As a result, UICs did not significantly differ between seasons in 24-hour collections and spot samples, whereas the spot UIC:UCC ratio differed by 21% (P<0.001) and reflected the lower iodine intake. In both study populations, the within- and between-person variabilities in urine volume, 24-hour UICs, and spot UICs were higher than the variability between seasons. Spot UIC may slightly overestimate the iodine intake in hot temperatures due to concentrated urine, and methods to correct for urine volume may be considered. Local seasonal differences in iodine intakes may also occur in some populations. This trial was registered at http://www.clinicaltrials.gov as NCT03215680.

MecA positive Staphylococcus spp. in bovine mastitis, milkers, milking environment, and the circulation of different MRSA clones at dairy cows farms in the Northeast region of Brazil

ABSTRACT: This study detected the presence and distribution of mecA in Staphylococcus spp. in the dairy production environment at farm level in Brazil. We analyzed 335 samples of mastitis cow milk, 15 samples of nostrils and hand swabs from milkers, 14 teat cup swabs, and 9 milking buckets swabs. Initially, the samples were subjected to microbiological analysis to detect Staphylococcus spp. and then S. aureus and mecA positive isolates were identified by PCR. All S. aureus isolates carrying the mecA genes were subjected to DNA macro-restriction analysis by Pulsed-Field Gel Electrophoresis (PFGE). The mecA gene was detected in 6/335 (1.78%) of mastitis cow milk, 5/15 (33.3%), and 5/15 (33.3%) of nostrils and hand swab, and 4/14 (28.5%) of the teat cup isolates. MRSA genotyping was performed by PFGE, a total of seven pulsotypes were grouped in two clusters. This study identified the occurrence and spread of MRSA at dairy environment of farms, and also the existence of distinct genetic profiles between isolates.

Identification of coagulase negative staphylococcal species from bovine mastitis in India.

Staphylococcal mastitis is a major cause of concern to the dairy industry in India and several countries worldwide. Though Staphylococcus aureus is the major cause, coagulase negative staphylococcal species (CoNS) are being increasingly reported in recent years. To investigate the incidence of coagulase negative staphylococcal species in bovine mastitis. Isolation of staphylococci was carried out from 237 milk samples of cows and She buffaloes with clinical and subclinical mastitis from different regions of Andhra Pradesh and Karnataka. CoNS isolates were identified by tube coagulase test using fresh rabbit plasma and coagulase gene PCR. We employed the biochemical test scheme published elsewhere previously for identification of the CoNS isolates up to species and subspecies levels. Seven representative isolates were identified by 16S rDNA sequencing to check the accuracy of biochemical test based identification. The CoNS constitute the majority of the staphylococcal isolates from mastitis (80/125, 64%) in this region. Using biochemical test scheme, the CoNS isolates from bovine mastitis were identified as S. cohnii sub sp. cohnii, S. simulans, S. capitis sub sp. capitis, S. cohnii sub sp. xylosus, and S. lugdunensis. The CoNS species S. schleiferi, S. haemolyticus, S. sciuri, S. xylosus, S. chromogenes, and Macrococcus epidermidis were identified by 16S rDNA sequencing. The 16S rDNA sequencing is the appropriate method for the identification of CoNS species. This study highlighted coagulase negative staphylococcal species as possible etiological agents of mastitis.

Quantification of mineral and toxic elements in milk of dairy cows by ICP-MS

Milk is an important component of mammalian metabolism which included micro and macronutrients. The objective of this study was to survey the concentrations of minerals and toxic elements in cow milk samples by inductively coupled plasma-mass (ICP-MS). In this study, it was analyzed the content of sodium, magnesium, potassium, calcium, titanium, chromium, manganese, iron, cobalt, nickel, copper, zinc, selenium, arsenic, cadmium, thallium, lead in cow milk samples (n=25) collected from Kahramanmaraş province of Turkey. The highest levels recorded for potassium, calcium and sodium with average concentrations of 1587.36, 1157.98, 643.34 mg L-1, respectively. The results revealed that there was no toxic element contamination in cow milk samples. Regular element analysis of cow's milk, which is the most widely used milk in dairy industry, is important for health.

A novel electrochemical aptasensor based on layer-by-layer assembly of DNA-Au@Ag conjugates for rapid detection of aflatoxin M1 in milk samples

Aflatoxin M1 (AFM1) is a common toxin in dairy products that causes acute and chronic human health disorders. Thus, the development of a rapid and accurate AFM1 detection method is of vital importance for food safety monitoring. This work was to develop a novel electrochemical aptasensor for sensitive and specific determination of AFM1. The dendritic-like nanostructure was formed on the gold electrode surface by layer-by-layer assembly of gold-silver core-shell nanoparticles modified with DNA conjugates. In the presence of AFM1, the specific recognition between AFM1 and Apt caused the disassociation of the DNA controlled dual Au@Ag conjugates from the surface of the electrode, causing less methylene blue to bind to the surface and weakening the electrochemical signal. The more AFM1 there is, the weaker the electrochemical signal. Transmission electron microscope results showed that the successfully synthesized Au@Ag nanoparticles exhibited a core-shell structure with Au as core and Ag as shell, and their average diameter was about 30 nm. Under optimal conditions, the electrochemical aptasensor showed a wide detection ranging from 0.05 ng mL-1 to 200 ng mL-1, and a low detection limit of 0.02 ng mL-1. Moreover, the proposed strategy has been successfully applied to the detection of AFM1 in cow, goat, and sheep milk samples with satisfactory recoveries ranging from 91.10% to 104.05%. This work can provide a novel rapid detection method for AFM1, and also provide a new sensing platform for the detection of other toxins.

Puerarin-coated gold nanoparticles (PUE-AuNPs) synthesized via green synthetic route: a new colorimetric probe for the detection of ciprofloxacin.

Puerarin-coated gold nanoparticles (PUE-AuNPs) synthesized via green synthetic route, a new colorimetric sensor, efficiently detected the ciprofloxacin (CP) in tap water and cow milk samples. The PUE-AuNPs were characterized by UV-visible, FTIR, AFM, and DLS techniques and were found to be spherical with an average size of approximately 19-20 nm. FTIR spectrum confirms that functional groups such as -OH, -C=O, -CO and -C=C were responsible for the reduction of gold (III) chloride trihydrate. These functional groups acted as capping agents to form AuNPs. The PUE-AuNPs sensor was proved to be selective and sensitive for the detection of CP through colorimetric method within the concentration of 1 to 1000 μM and the limit of detection was 51 μM. This colorimetric sensor is simple, cost-effective, and selective towards CP detection in environmental (tap water and milk) samples.

Evaluation of bacterial contaminants and heavy metals in cow and buffalo raw milk sold in Baghdad governorate

The purpose of this study was to investigate bacterial contamination and heavy metal concentrations in 80 samples of raw milk (cow:40 and buffalo:40) gathered from local markets in Baghdad, Iraq. The culture results were classified into ten categories: E. coli was 100% in each cows and buffaloes, Enterobacter Spp 23.75% (25% cow, 22.5% buffalo), Pseudomonas Spp 13.75% (15% cow,12.5% buffalo), Klebsiella Spp 15% (17.5% cow, 12.5% buffalo), Staphylococcus aureus 12.5% (15% cow, 10% buffalo), Staph. epidermidis 5% (for each cow and buffalo), Proteus spp. 10% (12.5% cows, 7.5% buffaloes), E. coli O157 15% (25% cow, 5% buffalo), Yersinia enterocolitica 3.75% (5% cow, 2.5% buffalo) and Salmonella 13.75% (25% cow 2.5% buffalo). The averages of heavy metals concentrations in cow milk samples were (0.62±0.25), (0.25±0.22), (0.31±0.20) and (21±2) mg/kg and in buffalo milk samples were (0.60±0.3), (0.33±0.15), (0.27±0.11) and (18±2.5) mg/kg for Lead (Pb), Cadmium (Cd), Copper (Cu) and Nickel (Ni) respectively. The high concentrations of pathogenic bacteria and metals found in the milk products is a sign of inadequate hygiene and sanitation during milking and post-milking operations, as well as excessive levels of heavy metal pollution in the environment which will affect meat and milk produced by these animals.

Interrelationship of milk acute-phase proteins and casein percentage in cows and buffaloes subclinical mastitis.

A total number of 62 clinically healthy dairy animals of three farms located in Kafr El Sheikh governorate, Egypt, were subjected to field screening surveys of subclinical mastitis (SCM) using California mastitis test (CMT). The obtained results revealed that 38.80% of quarter milk samples were positive to CMT. The most frequently major causative agents isolated from the positive CMT samples were Escherichia coli, Staphylococcus aureus, and environmental streptococcus spp. Acute-phase proteins (APPs), as immunological biomarkers for SCM, including milk serum amyloid A (mSAA) and haptoglobin (Hp) were measured using ELISA. A significant positive correlation was found between the severity of the mammary infection of cow's quarter milk samples represented in somatic cell count (SCC) and each of APPs and pH values. The correlation coefficient (R) between SCC and mSAA, Hp and pH were 0.54, 0.38 and 0.73, respectively. On the other hand, there was a significant negative correlation between casein percentage in milk of SCM cases, and each of APPs, pH and the presence of bacterial pathogens in the milk samples. The obtained results threw light on the inter-relationship between SCC, mSAA, pH value and casein percentage in milk of cows and buffalo suffered from SCM. The percentage of casein in milk is considered a significant accurate tool for diagnosis of SCM and this finding offers the farmers a cheap and fast selection for diagnosis of such disease. These results presented a specific structured view on the efficacy of different diagnostic tools of SCM in dairy herds.

Isolation and molecular detection of enterotoxigenic Staphylococcus aureus from raw milk of cows

Staphylococcus aureus is a microbe associated with human’s food poisoning. It is caused by isolates producing different heat stable enterotoxins which act as one of the most spread worldwide gastroenteritis causes. The goal of the current study was to assess the incidence of S. aureus (enterotoxigenic) in the raw milk samples of cows which were collected from various places of Baghdad city. Isolation and identification of S. aureus were done by conventional laboratory method and the diagnoses were confirmed by using molecular method RT-PCR to detect the specific gene of S. aureus nuc gene. In addition, we investigate the occurrence of some of Staphylococcal enterotoxins genes such as SEA, SEB, SEC, SED, and SEE. We found that 12/50 (24%) of the isolates were S. aureus and these isolates carried one or more of the enterotoxin genes such as enterotoxin type SEC 12/12 (100) followed by enterotoxin type SEA 7/12 (58.33%) and only 1/12 (8.33%) was type SED, while type SEB and SEE were negative 0%. Some of the isolates have genes that coded for two enterotoxins patterns such as SEA and SEC 7/12 (58.33%) and SEC and SED 1/12 (8.33%). Fewer of the isolates expressed only one genotype of enterotoxin gene like SEC 4/12 (33.3%). This study has proven that there was a high chance of occurrence for the enterotoxigenic S. aureus at the raw milk of cows in Baghdad city.

Impedance analysis of milk quality using functionalized polyamide textile-based sensor

Present paper demonstrates design and characterization of a textile based microfluidic chip sensor for the detection of milk adulteration through measuring the real part of the impedance and impedance phase angle. Polyamide (PA) based textile fabric was chemically functionalized with polyaniline and titanium dioxide nanoparticles (PANI/TiO2) nanocomposite and embedded in the microfluidic chip. Prototyping of microfluidic chip was performed by xurography and hot lamination using polyvinyl chloride foils. Morphological and chemical properties of fabricated textile-based PA-PANI/TiO2 chip sensor were evaluated by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). Quality of milk was accessed with fabricated textile sensor using cow and goat milk samples. The electrochemical impedance spectroscopy (EIS) was applied to detect the change in resistance and phase angle of pure and diluted milk. The developed PA-PANI/TiO2 chip sensor acted as a variable resistor that was able to identify adulterations and spoilage of the milk samples with sensitivity of 0.06 degrees of phase angle variation per % of water dilution. Our work promises that application of textile electronics could be efficiently exploited for food safety, point-of-care and environment monitoring applications.

The Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection.

Simple SummaryLiquid chromatography for determination of orotic acid (OAc) in milk of ewes and cows is presented. Acetonitrile was added to collected milk samples; then, the resulting mixture was centrifuged. Water was added to the obtained supernatant. Finally, the obtained solution was injected into two analytical columns. All analyses were performed at a column temperature of 35 °C. Suitable separation of OAc from endogenous species of milk can be achieved using the gradient elution program and UV detection. The current chromatographic procedure resulted in satisfactory precision, accuracy and sensitivity of OAc analyses in milk samples; OAc eluted at ca. 6.4 min. Our improved chromatographic method is suitable for routine determination of OAc in milk of sheep and cows.Ultra-fast liquid chromatography (UFLC) with a photodiode array detector (DAD) for simple and rapid determination of orotic acid (OAc) in milk of sheep and cows is described. Milk samples are treated with acetonitrile (1:1, v/v) and then centrifuged at 4 °C. To 1 mL of the obtained supernatant 9 mL of ultrapure water was added. Subsequently, 0.5–6 µL of the resulting solution was injected into the UFLC-DAD system. Separation and quantification of OAc in milk samples was achieved using two Kinetex C18 columns (1.7 µm, 150 mm × 2.1 mm, i.d., 100 Å; Phenomenex) fitted with a pre-column of 4 mm × 2 mm, i.d. (Phenomenex) containing C18 packing material. All separations were performed at a column temperature of 35 °C while the ambient temperature was 21–24 °C. Satisfactory separation of OAc from endogenous species of milk can be achieved using the binary gradient elution program and UV detection at wavelengths 278 nm. Our original procedure resulted in suitable separation and quantification of OAc in milk samples; OAc eluted at 6.44 ± 0.03 min. The total run time of OAc analysis (including re-equilibration) was 27 min. As expected, the OAc peak was absent from the blank when the proposed gradient elution program and UV detection at 278 nm was used. The average recoveries of OAc standards added to milk samples were satisfactory (96.7–105.3%). The low inter-and intra-assay coefficient of variation derived from the measurements of OAc in cow and ovine milk samples (i.e., 0.784%, 1.283% and 0.710%, 1.221%, respectively) and in O-Ac standards (i.e., 0.377% and 0.294%, respectively), as well as high recoveries of OAc added to ovine and cows’ milk (~100%) and the low detection (0.04 ng) and quantification (0.12 ng) limits point to satisfactory accuracy, precision and sensitivity of the reported method. OAc concentrations in ovine milk samples were within the range from 25 to 36 mg/L, while OAc levels in cows’ milk samples was found in the range of 32–36 mg/L. Our original procedure is suitable for routine quantification of OAc in milk of ewes and cows.

CSIWC RF sensor for micro‐fluidic non‐contact quality assessment of milk

In this work, the design of the SIW-based RF sensor is presented to assess the targeted quality of milk. In the present scenario, mainly two parameters, namely, purity and fat level are considered to assess the milk quality. Customized in-lab preparation of milk adulterant using low-grade vegetable oil, detergent, water, and glucose is carried out to analyze the practical milk adulteration scenario. Moreover, commonly consumed pure cow and buffalo milk samples are also collected to record the corresponding reference RF properties. A systematic approach is followed to develop the complete database related to actual milk samples with known fat percentages. Then accordingly, the proposed RF sensing mechanism is employed to identify the possible deviation in milk quality. The proposed work mainly employs the SIW-based non-contact RF sensing mechanism where the perturbation formulation is used to perform a qualitative assessment. Mathematical processing of the relative shift in the obtained S-parameters, corresponding to the variation in dielectric property of test samples at specified RF frequency, is carried out. Even though the work's main aim is a qualitative assessment of milk quality, dielectric property estimation of considered test samples is also performed to visualize their quantitative RF attributes. A compact, low-cost, non-contact, real-time, and micro-fluidic CSIWC RF sensor with high-quality factor and improved accuracy and sensitivity is designed.

Differences in post-translational modifications of proteins in milk from early and mid-lactation dairy cows as studied using total ion chromatograms from LC-ESI/MS

Liquid chromatography top-down mass-spectrometry combined with an in-house library with 5180 entries of known milk protein masses was used to identify common and rare isoforms of major milk proteins from individual cow's milk samples from early- and mid-lactating (EL and ML, respectively) cows. The milk samples were from 24 Danish Holstein-Friesians, with 12 cows each in EL and ML. A Latin square design with four periods was used to obtain one composite milk sample from each cow per period. The deconvoluted masses were used for identifying common and rare isoforms of the major milk proteins (αS1-CN, αS2-CN, β-CN, κ-CN, α-LA, and β-LG). αS1-CN splice variants were found in all samples, αS- and κ-CN isoforms differed between EL and ML. κ-CN post-translational modifications were found to relate to κ-CN genotype. This approach could identify 66 isoforms of major milk proteins, where 15 were of the αS-CNs and 39 from κ-CN.

Nuclear magnetic resonance spectroscopy to investigate the association between milk metabolites and udder quarter health status in dairy cows

Nuclear magnetic resonance spectroscopy was applied to investigate the association between milk metabolome and udder quarter health status in dairy cows. Mammary gland health status was defined by combining information provided by traditional somatic cell count (SCC) and differential SCC (DSCC), which expresses the percentage of neutrophils and lymphocytes over total SCC. Quarter milk samples were collected in triplicate (d 1 to 3) from 10 Simmental cows, 5 defined as cases and 5 defined as controls according to SCC levels at d 0. A total of 120 samples were collected and analyzed for bacteriology, milk composition, SCC, DSCC, and milk metabolome. Bacteriological analysis revealed the presence of mostly coagulase-negative staphylococci in quarter milk samples of cows defined as cases. Nuclear magnetic resonance spectra of all quarter samples were first analyzed using the unsupervised multivariate approach principal component analysis, which revealed a specific metabolomic fingerprint of each cow. Then, the supervised cross-validated orthogonal projections to latent structures discriminant analysis unquestionably showed that each cow could be very well identified according to its milk metabolomic fingerprint (accuracy = 95.8%). The comparison of 12 different models, built on bucketed 1-dimensional NOESY spectra (noesygppr1d, Bruker BioSpin) using different SCC and DSCC thresholds, corroborated the assumption of improved udder health status classification ability by joining information provided by both SCC and DSCC. Univariate analysis performed on the 34 quantitated metabolites revealed lower levels of riboflavin, galactose, galactose-1-phosphate, dimethylsulfone, carnitine, hippurate, orotate, lecithin, succinate, glucose, and lactose, and greater levels of lactate, phenylalanine, choline, acetate, O-acetylcarnitine, 2-oxoglutarate, and valine, in milk samples with high somatic cells. In the 5 cases, results of the udder quarter with the highest SCC compared with its symmetrical relative were in line with quarter-level findings. Our study suggests that increased SCC is associated with changes in milk metabolite fingerprint and highlights the potential use of different metabolites as novel indicators of udder health status and milk quality.

Within-herd prevalence threshold for the detection of Mycobacterium avium ssp. paratuberculosis antibody–positive dairy herds using pooled milk samples: A field study

Herd-level diagnosis of paratuberculosis using a pool-milk ELISA (pool size: n ≤ 50) is a novel, economical, and convenient method to identify blood serological Mycobacterium avium ssp. paratuberculosis (MAP) antibody-positive herds. To date, the diagnostic performance of the pool-milk ELISA has been described only under laboratory conditions where herd prevalence was simulated by the preparation of milk pools consisting of milk samples of cows with a known MAP status determined by fecal culture. In our observational study, test performance under field conditions was studied using pooled milk and individual blood samples. A total of 486 herds within the MAP prevalence reduction program of Lower Saxony, from which pooled milk and individual blood ELISA results were available, were assigned to this study. Data were analyzed for the period between January 1 and December 31, 2018, the first year after herd testing became obligatory in this federal state of Germany. To evaluate whether pooled milk samples reliably distinguish between herds with a MAP-apparent blood serological within-herd prevalence (MAP-Ab-WHPapp) ≥5% and herds with a MAP-Ab-WHPapp <5%, the distribution of the MAP-Ab-WHPapp was compared between pool-positive and pool-negative herds. The MAP-Ab-WHPapp was 3.4% (median; 95% confidence interval = 0-11.4%) in pool-positive herds and 1.2% (median; 95% confidence interval = 0-6.4%) in pool-negative herds. Only 10.8% (n = 12) of the pool sample-negative herds had a MAP-Ab-WHPapp ≥5% and were therefore false negatives, given the aims of the MAP prevalence reduction program. Hence, the pool-milk sampling strategy seems well suited to distinguish between herds with a MAP-Ab-WHPapp ≥ 5% and herds with a MAP-Ab-WHPapp <5% since only 10% of serum MAP-ELISA positive herds were missed. Employing a logistic regression model, we estimated that the minimum blood serological MAP-Ab-WHPapp to detect a pool-positive herd with a probability of 95% was 8%, which fits well with the aim of the MAP prevalence reduction program to focus on herds with a MAP-Ab-WHPapp of ≥5%. Despite the limitations of the control approach, which include milk pool sample collection and a low sensitivity of the ELISA used in milk pools and serum samples, the aims of the MAP prevalence reduction program can be achieved. The results of these field data support that pool-milk sample ELISA is a useful, economical, and low labor-intensive tool to identify herds seropositive for MAP in a MAP prevalence reduction program.

βeta-lactam and Tetracycline Antibiotic Residues in Cow Milk in the Constantine region, Algeria

The aim of the present study was to investigate the βeta-lactam and tetracycline antibiotic residues in cow milk samples. A total of 122 samples of cow milk were collected from raw milk collectors (109 samples) and from a reconstituted pasteurized milk sales clerk (13 samples) in the Constantine region, Algeria and examined using the ΒetaStar Combo screening kit (Neogen, USA). Results indicates that 13 samples (10.66%) were positive for antibiotics residues: 12 (9.84%) for βeta-lactams (ten (8.20%) raw and two (1.64%) pasteurized milk samples) and only one (0.82%) for tetracyclines in a raw milk sample. It is evident that the Algerian consumer is not sheltered from the danger of antibiotic residues in milk and these inhibitor residues should constitute a constant concern for the dairy industry in Algeria. A control programme should be established.

Application of the Actiphage® Assay to Detect Viable Mycobacterium avium subsp. paratuberculosis Cells in Fresh Sheep and Goat Milk and Previously Frozen Milk and In-Line Milk Filters.

Mycobacterium avium subsp. paratuberculosis (MAP) is a well-known causative agent of paratuberculosis, a chronic infectious granulomatous enteritis of ruminants contributing to significant economic losses worldwide. Current conventional diagnostic tools are far from being sufficient to manage and control this disease. Therefore, increased attention has been paid to alternative approaches including phage-based assays employing lytic bacteriophage D29 to detect MAP cells. The aim of the present study was to assess the applicability and efficiency of the recently developed phage-based kit termed Actiphage® combined with IS900 real-time PCR (qPCR) for rapid detection and quantification of viable MAP in milk samples. We demonstrated that Actiphage® in combination with IS900 qPCR allows for rapid and sensitive detection and identification of viable MAP in milk samples with a limit of detection of 1 MAP per 50 ml milk. Using this method, the presence of viable MAP cells was successfully determined in 30.77% of fresh goat, sheep and cow milk samples originating from paratuberculosis-affected herds. We further used Actiphage assay to define the time-lapse aspect of testing naturally contaminated milk and milk filters frozen for various lengths of time by phage-based techniques. Viable MAP was detected in 13.04% of frozen milk samples and 28.57% of frozen milk filters using Actiphage-qPCR. The results suggest the ability to detect viable MAP in these samples following freezing for more than 1 year. The obtained results support the views of the beneficial role of this technology in the control or monitoring of paratuberculosis.

Antimicrobial resistance of Staphylococcus aureus strains isolated from cow raw milk samples from Albania and Serbia

This research aimed to determine the prevalence rate of antimicrobial resistance (AMR) of Staphylococcus aureus isolated from raw cow milk samples in Albania and Serbia. A total of 100 raw milk samples, 50 from Albania and 50 from Serbia, have been collected from randomly selected cattle farms. Twelve samples (12%) were positive for S. aureus, five of those originating from Albania (41.66%) and seven from Serbia (58.33%). Resistance of isolated S. aureus strains to ampicillin in milk samples from Albania (36.7%) and Serbia (34.1%) was the most common. Some of S. aureus strains from milk samples from Albania were resistant to tetracycline (16.9%), while isolates from milk samples from Serbia were more resistant towards oxacillin (18%). The recorded resistance towards erythromycin (13.2%; 13.1%), and sulfatrim (7.6%; 6.9%) was similar between both milk samples, respectively. The obtained results have shown higher resistance of S. aureus strains towards cephalothin in milk samples from Serbia (3.9%) compared to milk samples from Albania (1.6%), respectively. All isolates tested for antibiotic sensitivity were susceptible to methicillin, vancomycin, chloramphenicol, and ciprofloxacin. The obtained results regarding the minimum inhibitory concentrations (MICs) of antibiotics indicated that isolates were resistant to tetracycline, oxacillin, erythromycin, and ampicillin, respectively. Results of our investigation have shown that the found antibiotics residues are related to the usage of erythromycin (72.6% and 73.9%), followed by ampicillin (70.4% and 71.2%) while residues of ciprofloxacin in the analysed milk samples was not registered.

Comparison of organic and deep eutectic solvents based dispersive liquid-liquid microextraction for the analysis of phytosterols in cow milk combined with high-performance liquid chromatography-ultraviolet detector.

In the present work, a dispersive liquid-liquid microextraction approach has been developed for extraction of four phytosterols (stigmasterol, β-sitosterol, campesterol, and brassicasterol) from cow milk samples using organic and deep eutectic solvents and the results were critically compared. The extracted analytes were determined using high performance liquid chromatography. In the developed method, carbon tetrachloride and choline chloride:p-chlorophenol deep eutectic solvent were selected to use as the best extraction solvent. Effective parameters and validation data were studied for both methods, independently. Under optimum conditions, limits of detection and quantification were within the ranges of 0.3-0.9 and 1.0-3.0ng/mL for organic solvent based dispersive liquid-liquid microextraction and 0.09-0.32 and 0.3-1.0ng/mL for deep eutectic solvent based dispersive liquid-liquid microextraction, respectively. Good coefficient of determinations and relative standard deviations obtained for the methods were ≥0.994 and ≤7.6%, respectively. The introduced method was performed on different milk samples for the determination of target analytes using both solvents and the results were analyzed statistically by the t-test.

A1 and A2 milk caseins-comparative FTIR and spectroflourimetry analysis

Around 35% of the total caseins are β-caseins, which are further classified as A1 β-caseins and A2 β-caseins, based on differences in the amino acid composition of both. A2 is the wild type genetic variant of β-casein while A1 is the mutant. The present study aimed at the isolation of A1 and A2 casein from different cow milk sources and its characterization by using simple chemical techniques, viz. FTIR and spectrofluorimetry. The commercial milk sample from Bos indicus (Gir) (A2) was obtained from Bombay Panjrapole, Mumbai and two commercially available packaged cow milk samples (pasteurized, skimmed) namely from Gokul and Mother Dairy (A1) were also obtained for comparison analysis from the local market. The isolation of casein was performed by standard method and analyzed using SDS-PAGE, FTIR and spectrofluorimetry. There was evidence that the A2 milk lacked histidine and rich in aromatic amino acids like tryptophan using FTIR and spectrofluorimetry techniques.