不同生鲜水牛乳滴定酸度的分析

按GB 4789.18—2010《乳与乳制品检验》标准对云南省主要奶水牛养殖小区、合作社同一季节26户养殖户的228个水牛生乳及相同来源不同月份的655个水牛生乳的酸度进行滴定,同时进行酒精检测和掺碱试验,并对日粮组成、饲喂模式及胎次和泌乳天数进行普查。同一季节不同来源的228个水牛生乳滴定酸度在10~13°T的占83.8%;相同来源不同月份的655个水牛生乳样品的滴定酸度在10~13°T的占86.4%;总的883个生鲜水牛乳滴定酸度为10~13°T的占85.7%,大于14°T占10.6%,小于10°T仅占3.6%。     

Genotypic analysis of nontuberculous mycobacteria isolated from raw milk and human cases in Wisconsin

Nontuberculous mycobacteria (NTM) compose a group of mycobacteria that do not belong to the Mycobacterium tuberculosis complex group. They are frequently isolated from environmental samples such as water, soil, and, to a lesser extent, food samples. Isolates of NTM represent a major health threat to humans worldwide, especially those who have asthma or are immunocompromised. Human disease is acquired from environmental exposures and through consumption of NTM-contaminated food. The most common clinical manifestation of NTM disease in human is lung disease, but lymphatic, skin and soft tissue, and disseminated disease are also important. The main objective of the current study was to profile the farm-level contamination of cow milk with NTM by examining milk filters and bulk tank milk samples. Five different NTM species were isolated in one dairy herd in Wisconsin, with confirmed 16S rRNA genotypes including Mycobacterium fortuitum, Mycobacterium avium ssp. hominissuis, Mycobacterium abscessus, Mycobacterium simiae, and Mycobacterium avium ssp. paratuberculosis (Mycobacterium paratuberculosis). In tank milk samples, M. fortuitum was the predominant species in 48% of the samples, whereas M. chelonae/abscessus and M. fortuitum were the only 2 species obtained from 77 and 23% of the examined filters, respectively. Surprisingly, M. avium ssp. hominissuis, M. paratuberculosis, and M. simiae were isolated from 16.7, 10.4, and 4% of the examined milk samples, respectively, but not from milk filters. Interestingly, NTM isolates from human clinical cases in Wisconsin clustered very closely with those from milk samples. These findings suggest that the problem of NTM contamination is underestimated in dairy herds and could contribute to human infections with NTM. Overall, the study validates the use of bulk tank samples rather than milk filters to assess contamination of milk with NTM. Nontuberculous mycobacteria represent one type of pathogens that extensively contaminate raw milk at the farm level. The significance of our research is in evaluating the existence of NTM at the farm level and identifying a simple approach to examine the potential milk contamination with NTM members using tank milk or milk filters from dairy operations. In addition, we attempted to examine the potential link between NTM isolates found in the farm to those circulating in humans in Wisconsin.     

The prevalence and characterization of Bacillus cereus isolated from raw and pasteurized buffalo milk in southwestern China

Bacillus cereus is an important food-borne pathogenic bacteria and a putrid microorganism in the dairy industry. Raw and pasteurized buffalo milk play important roles in the dairy market in southwestern China. However, the reports on the prevalence and characterization of B. cereus strains isolated from the above sources are lacking. In this study, 150 raw buffalo milk samples and 300 pasteurized buffalo milk samples were collected from 3 provinces in southwestern China. The genotype, virulence gene distribution, antibiotic resistance, and biofilm-forming ability of isolates were analyzed. Ninety-six B. cereus strains were isolated and identified: 50 isolates (33.3%) from buffalo raw milk and 46 isolates (15.3%) from pasteurized buffalo milk. These strains were classified into 41 sequence types (ST) and 5 groups, of which ST857 was the predominant ST. The detection rates of virulence genes nheABC cluster, hblACD cluster, cytK, bceT, entFM, hlyII, and cesB were 89.6%, 13.5%, 64.6%, 71.9%, 84.4%, 62.5%, and 6.25%, respectively. The antimicrobial susceptibility testing showed that more than 90% of the isolates were susceptible to gentamicin, chloramphenicol, ciprofloxacin, erythromycin, vancomycin, and tetracycline, as well as resistant to ampicillin, cefepime, oxacillin, and rifampin. The results of biomass biofilm evaluation of the isolates on the stainless-steel tube showed that the optical density values at a wavelength of 595 nm of all strains in group I were greater than 1, with the strongest overall biofilm-forming ability among 5 groups, and the overall biofilm-forming ability of group III was the weakest. There was a relationship between the biofilm-forming ability and phylogenetic relationship of B. cereus strains. Taken together, our findings are the first to report the contamination situation and characterization of B. cereus isolated from raw and pasteurized buffalo milk in southwestern China as well as indicate the potential risk posed by this pathogen to dairy industry and public health.     

Characterization of the major whey proteins from milk of Mediterranean water buffalo (Bubalus bubalis)

In this work, the whey protein fractions from 120 Mediterranean water buffalo individual milks were analysed by microchip electrophoresis (MCE), reverse-phase high-performance liquid chromatography (RP-HPLC) and mass spectrometry (ESI-MS). Validation procedures were carried out for both MCE and HPLC. The chromatographic analysis allowed the complete separation of the whey protein fractions, resulting in a well-defined peak structure; the adopted RP-HPLC and ESI-MS protocols provided identification of β-lactoglobulin (18,266 Da), α-lactoalbumin (14,236 Da) and serum albumin (66,397 Da). The calculated mean concentrations were 4.04 g/l, 2.45 g/l and 0.35 g/l, respectively.     

Detection of buffalo milk adulteration with cow milk by capillary electrophoresis analysis

The addition of cow milk during the production of buffalo mozzarella is a common fraud in dairy industries because of the lower price and greater availability of cow milk throughout the year. The aim of this study was to develop a new, rapid, and robust capillary electrophoresis method for detecting and quantifying cow milk in buffalo milk by exploiting cow α-lactalbumin as a marker of adulteration. In particular, a linear calibration curve was generated, using a training set of calibrators consisting of 7 series of 17 buffalo/bovine whey mixtures, obtained after casein precipitation, with increasing percentages of cow whey. The capillary electrophoresis method showed high linearity (R² = 0.968), repeatability [relative standard deviation (RSD) = 2.11, 3.02, 4.38, and 1.18%, respectively for 5, 10, 20, and 50% of buffalo/bovine whey mixtures], and intermediate precision (RSD = 2.18, 2.49, 5.09, and 3.19%, respectively, for 5, 10, 20, and 50% buffalo/bovine whey mixtures). Moreover, the minimum amount of detectable fraudulent cow milk was 1%, and the limit of quantification was 3.1%.     

Characterisation and comparison of khoa prepared from camel milk with that from cow and buffalo milk

Khoa, a heat‐concentrated milk product, is used as a base material for the manufacture of many popular sweets. The comparison was made between various chemical compositions and characteristics of the khoa prepared from the camel milk with that prepared from the cow and the buffalo milk samples. The khoa prepared from the camel milk had the higher moisture, ash, acidity, soluble nitrogen, free fatty acids and peroxide value, but lower in fat, protein and lactose contents than that prepared from the cow and buffalo milk samples. There was no significant (P > 0.05) difference in 5‐hydroxymethyl furfural between khoa samples prepared from the three milks.     

A signature protein-based method to distinguish Mediterranean water buffalo and foreign breed milk

A novel genetic variant at the α_s1-casein locus of water buffalo (WB), 8-residue shorter than its wild-type has been found and sequenced. The internal deletion of the peptide E³⁵KVNELsT⁴² was confirmed by the isolation of the junction peptide. The 8-residue deletion mutant has a molecular weight that is 919 Da less than that of the wild-type. The novel isoform with a unique f35-42 deletion could be the result of the skipping of exon 6, generating an exon 6-deleted variant of α_s1-casein. The wild-type and its shortened α_s1-casein forms were found to co-exist in many individual milk samples. In contrast, the 8-residue, internally deleted α_s1-casein variant did not occur in water buffaloes of the Mediterranean breed reared in Italy. Wild-type α_s1-casein has 6 to 8 phosphate groups (P) while the internally deleted form 6 and 7P per molecule.     

Mineral content of longissimus dorsi thoracis from water buffalo and Zebu-influenced cattle at four comparative ages

The longissimus dorsi thoracis (LDT) of 64 water buffalo and 68 Zebu-influenced cattle was used to determine the mineral content variation (mg/100g of fresh tissue) according to species, age (7, 17, 19 and 24 months-of-age, MOA) and gender (steers and bulls). Buffalo LDT had higher concentrations of K, Mn, Fe, Zn and Cu at weaning (P<0.05). Phosphorous content was higher (P<0.05) at post-weaning ages. Mineral profile was affected by age without clear trends; the Na and K contents increased at 24 MOA. The age×species, age×gender and species×gender interactions affected (P<0.05) Fe content that ranged from 1.74 to 2.56mg/100g fresh tissue. Meat from both species contributes with more of 25% of K, P, Fe and Zn of the daily requirement for an adult.     

Short communication: Detection of undeclared presence of bovine milk in buffalo yogurt

The authenticity of buffalo (Bubalus bubalis) dairy products is a focal issue, considering the increasing demand for buffalo milk products. Therefore, the aim of this study was to investigate the undeclared presence of bovine (Bos taurus) milk in buffalo yogurt, to understand which risk factors might make the product vulnerable to fraud. Real-time PCR assay showed the undeclared presence of bovine DNA in addition to buffalo DNA in 18 of 72 samples. Given the widespread lack of data on the presence of undeclared milk species in buffalo dairy products, the study provides a significant insight into the incidence of fraud in the buffalo dairy field. The data from this study could help improve the analysis of food safety risks along the buffalo milk supply chain and in the dairy processing industry, perceived as being highly vulnerable to food fraud, and prioritize target areas for food policy making to steer and enforce European food fraud regulations.     

水牛乳中可培养乳酸菌多样性分析

采用传统分离培养方法,从三品杂交生水牛奶混合样品中,分离出105株乳酸菌,通过形态、生理生化、API细菌鉴定系统及16S rDNA基因序列分析方法对各菌株属种进行鉴定。16S rRNA序列分析结果显示,105株菌共分为5个属8个种,呈现较为丰富的乳酸菌多样性,具体数量分布为乳酸乳球菌21株,植物乳杆菌19株,格氏乳球菌17株,乳明串珠菌13株,食窦魏斯氏菌11株,肠膜明串珠菌8株,类肠膜魏斯氏菌6株,嗜热链球菌5株,糊精乳杆菌5株。由此可知,水牛乳中可培养乳酸菌优势菌群的主次关系为：乳酸乳球菌（Lactococcus lactis）＞植物乳杆菌（Lactobacillus plantaru）＞格氏乳球菌（Lactococcus garvieae）＞乳明串珠菌（Leuconostoc lactis）＞食窦魏斯氏菌（Weissella cibaria）,此为后续开发水牛乳中优势乳酸菌资源提供了良好的理论基础。     

基于双向电泳和质谱联用技术的水牛乳源酪蛋白的研究

利用双向电泳和基质辅助激光解吸电离飞行时间质谱(MALDI-MS)联用技术对水牛乳酪蛋白与其他乳源蛋白差异性进行了研究。根据Image Master 2D Platinum图像分析软件对不同乳源酪蛋白的双向电泳(2-DE)图谱进行蛋白斑点的匹配分析,获得21个存在于水牛奶中,主要分布在低丰度蛋白区的差异蛋白点,经质谱分析,得到4个属于水牛奶酪蛋白的主要组分,另外发现两个与水牛奶中的蛋白有较高同源性的新组分。     

高效液相色谱-三重四级杆质谱联用法检测荷斯坦和水牛原料乳中磺胺类药物残留

目的分析目前我国南方地区荷斯坦原料乳和水牛原料乳中磺胺类药物的残留情况。方法采集浙江和广西地区荷斯坦牛牧场和水牛牧场的120份原料乳,采用高效液相色谱-三重四级杆质谱联用法检测其中的磺胺类药物残留。结果方法的检出限和回收率分别为0.05~0.10μg/L和73.2%~108.3%。水牛乳中磺胺类药物的残留量低于6.67μg/L,荷斯坦牛乳中磺胺类药物的残留量低于7.88μg/L。结论高效液相色谱-三重四级杆质谱联用法检测牛乳中磺胺类药物残留的结果准确可靠,我国南方地区的荷斯坦原料乳和水牛原料乳总体上处于安全范围。     

Heavy metal concentrations in raw milk collected from different regions of Samsun,Turkey

In this study, 144 raw milk samples were analysed for heavy metal contamination derived from emissions from industrial operations in Tekkekoy, Samsun, Turkey. Cu, Fe, Zn, Cr, Ni, Cd, As and Pb levels in samples were determined by inductively coupled plasma mass spectrometry (ICP‐MS). The average amounts of copper, chromium, iron, zinc, nickel, cadmium, arsenic and lead were determined as 1.130, 0.441, 12.920, 0.032, 0.483, 0.006, 0.003 and 0.004 mg/kg, respectively. It was determined that the summer period has the highest levels for copper, lead and cadmium. The highest contents of arsenic and copper were found at the two industrial regions that were close to Black Sea, described as ‘1st’ and ‘2nd’ region. Whereas the 3rd and 4th regions that were far from an industrial zone and also from the Black Sea, nickel, lead and chromium had the highest levels.     

金黄色葡萄球菌在生乳与灭菌乳中生长状况分析

目的 了解并比较金黄色葡萄球菌(S.aureus)在生乳与灭菌乳中的生长状况.方法 采用GB/T4789.37-2008<食品微生物学检验金黄色葡萄球菌检验>,测定不同温度下金黄色葡萄球菌在生乳和灭菌乳中的生长情况,利用MicroFit 1.0软件对所测数据进行分析,利用SPSS统计软件对该菌在生乳与灭菌乳中生长参数进行统计分析.结果 金黄色葡萄球菌在生乳中的最大生长速率明显低于在灭菌乳中的最大生长速率,二者的迟滞期、代时无明显差异.结论 金黄色葡萄球菌在生乳与灭菌乳中的生长情况差异显著.有统计学意义(P<0.05).     

Genome-wide association studies to identify quantitative trait loci affecting milk production traits in water buffalo

Water buffalo is the second largest resource of milk supply around the world, and it is well known for its distinctive milk quality in terms of fat, protein, lactose, vitamin, and mineral contents. Understanding the genetic architecture of milk production traits is important for future improvement by the buffalo breeding industry. The advance of genome-wide association studies (GWAS) provides an opportunity to identify potential genetic variants affecting important economical traits. In the present study, GWAS was performed for 489 buffaloes with 1,424 lactation records using the 90K Affymetrix Buffalo SNP Array (Affymetrix/Thermo Fisher Scientific, Santa Clara, CA). Collectively, 4 candidate single nucleotide polymorphisms (SNP) in 2 genomic regions were found to associate with buffalo milk production traits. One region affecting milk fat and protein percentage was located on the equivalent of Bos taurus autosome (BTA)3, spanning 43.3 to 43.8 Mb, which harbored the most likely candidate genes MFSD14A, SLC35A3, and PALMD. The other region on the equivalent of BTA14 at 66.5 to 67.0 Mb contained candidate genes RGS22 and VPS13B and influenced buffalo total milk yield, fat yield, and protein yield. Interestingly, both of the regions were reported to have quantitative trait loci affecting milk performance in dairy cattle. Furthermore, we suggest that buffaloes with the C allele at AX-85148558 and AX-85073877 loci and the G allele at AX-85106096 locus can be selected to improve milk fat yield in this buffalo-breeding program. Meanwhile, the G allele at AX-85063131 locus can be used as the favorable allele for improving milk protein percentage. Genomic prediction showed that the reliability of genomic estimated breeding values (GEBV) of 6 milk production traits ranged from 0.06 to 0.22, and the correlation between estimated breeding values and GEBV ranged from 0.23 to 0.35. These findings provide useful information to understand the genetic basis of buffalo milk properties and may play a role in accelerating buffalo breeding programs using genomic approaches.     

Effectiveness of poll stunning water buffalo with captive bolt guns

Practical experience has indicated that shooting water buffalo with a captive bolt gun in the front of the head does not always produce an effective stun. Slaughtermen have been claiming that the poll position is more reliable, but under present EU regulations this shooting position is not allowed for domesticated bovines. This study examined the effectiveness of shooting water buffalo with captive bolt guns in the poll region. The depth of concussion was assessed in 30 water buffalo from physical collapse, presence or absence of corneal and palpebral reflexes, normal rhythmic breathing, eyeball rotation, nystagmus and whether the animal was re-shot. All except one animal collapsed immediately. The one animal that failed to collapse was not breathing. The prevalence of a shallow depth of concussion in the poll shot animals was 53%. Two had been shot through the spinal cord, and with the exception of those plus one other in which the bolt did not penetrate the brain, the other animals were considered to be adequately stunned. Post mortem examination of the brain showed that 79% of the under thirty month old animals were shot in the midbrain or brain stem, but these regions had to be avoided in over thirty month old animals because of the risk of damaging the brain stem sample needed for BSE testing. In addition, three water buffalo were shot in the crown position and one in the front of the head. The frontal position was ineffective as the animal did not collapse, and all animals shot in the crown position resumed breathing shortly after shooting, indicating a shallow depth of concussion. It is concluded that poll shooting can be effective in water buffalo, but it produces a shallow depth of concussion compared with frontal shooting in cattle. It requires accurate placement of the gun to ensure that buffalo are not shot through the spinal cord instead of the brain. Vigilance is needed in ensuring that the animals are stuck promptly so that none recover consciousness.     

水牛奶乳清蛋白制备抗氧化活性肽工艺的研究

实验是以水牛奶为原料,分离纯化后得到乳清蛋白。利用碱性蛋白酶、中性蛋白酶、胰蛋白酶、木瓜蛋白酶和胃蛋白酶5种不同的蛋白酶对水牛奶乳清蛋白酶解以制备抗氧化活性多肽。酶筛选结果显示,中性蛋白酶是最适宜酶解水牛奶乳清蛋白制备抗氧化活性肽,其酶解液的还原能力和DPPH自由基清除率较其他4种酶高。探讨酶解反应时pH、温度、时间、酶浓度对酶解反应的水解度、酶解液的还原能力和DPPH自由基的清除率的影响,在单因素试验基础上,采用响应面法对酶解工艺进行优化。结果表明,中性蛋白酶酶解乳清蛋白的最佳工艺参数为:pH为7.4,温度为50.5℃,酶与底物浓度比为2.1%,酶解时间5.0h,此时2mg/mL酶解物的DPPH自由基清除率为32.58%。实测结果与预测值吻合效果良好。     

双抗体夹心酶联免疫法测定水牛初乳中的IgG

为快速、准确地测定水牛初乳中IgG活性质量浓度,建立了双抗体夹心酶联免疫分析法(ELISA)。结果表明,IgG活性质量浓度的对数值与吸光值存在很好的线性关系,其标准曲线方程式为γ=-0.2907x2+2.0819x+0.8689,相关系数R2=0.9937,线性范围在7.8～1000ng/mL,平均回收率在84.40%～96.71%之间,灵敏度高、重现性好,可用于水牛初乳及其制品中IgG活性质量浓度的检测。