Conventional identification of Streptococcus uberis isolated from bovine mastitis in Argentinean dairy herds

The objective of this study was to evaluate a conventional scheme for identifying Streptococcus uberis strains isolated from bovine mastitis. Seventy-five gram-positive, catalase-negative cocci were collected from cows with mastitis from 19 dairy herds located in the east-central region of Argentina. Five American Type Culture Collection strains and bovine isolates were identified by the API 20 Strep system and by restriction fragment length polymorphism analysis of 16S rDNA. A conventional scheme based on 11 biochemical tests was selected for identification of Strep. uberis strains: the Christie-Atkins-Munch-Petersen reaction; hydrolysis of Arg, esculin, and sodium hippurate; growth in inulin, mannitol, raffinose, salicin, and sorbitol; and growth at 45°C and in 6.5% NaCl. Reference strains and 25 bovine isolates were classified accurately to the species level by the conventional scheme in a blind assay. Each reference strain and each bovine isolate were identified as belonging to the same species following these 3 methods. The remaining 50 isolates identified as Strep. uberis by the API 20 Strep system and 16S rDNA RFLP were assayed by the conventional scheme. This scheme correctly identified 47 (94%) of 50 isolates as Strep. uberis by comparing their biochemical profile with that of the reference strain. Three (6%) of the 50 isolates were classified as Strep. uberis by the API 20 Strep system and by 16S rDNA RFLP and were identified as Enterococcus faecalis by the conventional scheme. Thirty percent of the Strep. uberis strains showed biochemical profiles identical to the Strep. uberis American Type Culture Collection 27958 strain. Seventy percent of the Strep. uberis strains demonstrated variability compared with the reference strain, resulting in 19 different biochemical profiles. The conventional scheme proposed in this study resulted in a relatively low number of misidentifications and could biochemically identify not only typical, but also atypical Strep. uberis strains. This conventional scheme can be considered an adequate method for identifying Strep. uberis strains isolated from bovine mastitis because of its affordable cost in developing countries, and it may contribute to determining the frequency of isolation of Strep. uberis strains in Argentinean dairy herds.     

Diversity of Salmonella isolates using serotyping and multilocus sequence typing

One hundred and twenty-one Salmonella isolates were obtained from food, feed, and live chicken samples derived from 13 countries or regions. In this study, their subtypes were evaluated by serotyping and multilocus sequence typing (MLST), and their genetic profiles were also characterized. It was demonstrated by serotyping on these isolates that 36 various serovars were obtained in this study, of which three serotypes S. Babelsberg, S. Fresno, and S. II were first found in mainland China. Based on Simpson’s index of diversity, the serotyping method had a 0.943 discriminatory power. Meanwhile, there were a total of 42 unique sequence types (STs) characterized by MLST, and the discriminatory power of MLST (D = 0.947) was close to that of the serotyping method. In MLST, hisD revealed the highest levels of nucleotide diversity. In addition, ST-92 was the most common ST represented by 16 Salmonella isolates, followed by ST-367 which was represented by 14 isolates. Seven new alleles were identified, which were associated with other alleles and resulted in the assignment of nine new STs. It was concluded from the results that MLST was generally associated with serotype, but not associated with the epidemiological source of the samples, and antimicrobial resistance patterns.     

Relationship between milk lactoferrin and etiological agent in the mastitic bovine mammary gland

Bovine mastitis is one of the most deleterious diseases for dairy herds and is mainly caused by contagious and environmental bacterial pathogens. Among contagious bacteria, Staphylococcus aureus is the most prevalent, whereas the main environmental mastitis pathogens are Streptococcus uberis and Escherichia coli. Bovine lactoferrin (bLF) is an approximately 80-kDa glycoprotein present in milk that participates in the innate response of the mammary gland against bacterial infection. The objectives of the current study were to analyze potential changes in bLF milk concentration, which would constitute a response of the mammary gland toward mastitis induced by different etiologic agents, and to evaluate a possible relation between this response and pathogen susceptibility to bLF. Microbiology analysis and bLF quantification in milk from different bovine mammary gland quarters were performed. Infected quarters presented greater concentrations of bLF compared with those from microbiologically negative quarters. Analysis of individual pathogen contributions showed that most of this increase was attributable to Strep. uberis intra-mammary infection. The ability of mammary gland cells to synthesize bLF in response to Strep. uberis challenge was demonstrated by immunodetection of the protein in in vitro infection experiments. Susceptibility of Strep. uberis, E. coli, and Staph. aureus to the antimicrobial activity of bLF was determined by growth inhibition assays conducted with 4 different isolates of each species. Whereas Staph. aureus and E. coli were shown to be susceptible to this protein, Strep. uberis appeared to be resistant to the antimicrobial activity of bLF. Molecular typing of the 4 Strep. uberis isolates used throughout this study showed that this result was representative of the species and not exclusive of a particular strain. Results presented herein suggest that different bacteria species may elicit different mammary gland responses mediated by bLF secretion and that Strep. uberis has probably adapted to this immune reaction by developing resistance to bLF inhibitory action.     

Field observations on the variation of Streptococcus uberis populations in a pasture-based dairy farm

Microbiological and molecular tools were used to monitor Streptococcus uberis populations on farm tracks and paddocks on a dairy farm during different seasons of a year to identify and profile potential environmental niches of Strep. uberis in a pasture-based dairying system. Farm tracks of high or low cow traffic were sampled every 2 wk for an entire year and Strep. uberis numbers were enumerated from a selective medium. During each season of the year, paddocks were sampled for the presence of Strep. uberis before and after grazing by dairy cows. Farm tracks of high cow traffic generally had greater concentrations of Strep. uberis isolated compared with tracks with less cow traffic, but there was also significant variation in the concentrations of Strep. uberis contamination among seasons, being highest in winter and lowest in summer. The bacterium was detected in paddocks only after cow grazing had occurred, but the bacteria could still be detected in soil for up to 2 wk following grazing in winter. Multilocus sequence typing showed great heterogeneity, with some commonality between farm track and milk isolates, which may help explain cow-to-environment or environment-to-cow transmission of the bacterium in the dairy setting.     

Molecular epidemiology of recurrent clinical mastitis due to Streptococcus uberis: Evidence of both an environmental source and recurring infection with the same strain

This study was undertaken because clinicians and farmers have observed that a considerable number of cows diagnosed with Streptococcus uberis mastitis have recurrences of mastitis in the same or a different quarter. The study was an attempt to answer whether these recurring cases were due to treatment failure (in which case a search would have begun for a better treatment for Strep. uberis mastitis) or due to reinfection with a different strain of Strep. uberis. Using pulsed-field gel electrophoresis (PFGE), we determined that the majority of recurrences (20 of 27) were caused by a new strain of Strep. uberis, indicating that treatment of the initial infection had been successful. A small number of recurrences (5 of 27) were caused by the initial strain, indicating persistence. The remaining 2 recurrences occurred in a new quarter but with the initial strain of Strep. uberis, indicating either spread between quarters or reactivation of a previous subclinical infection. Analysis of the PFGE profiles failed to reveal any strain-specific propensity to persist, because strains causing recurrences occurred in most of the major clusters.     

婴幼儿配方羊乳粉生产环节蜡样芽孢杆菌分离株多位点序列分型分析

为探讨我国婴幼儿配方羊乳粉生产环节阳性分离株的遗传多样性,并将其基因序列与已报道的乳源分离株的基因序列进行比对,确立其特征序列,为婴幼儿配方食品生产体系溯源提供依据,为探究其致病机理和有效防控提供参考依据。实验选取glpF、gmk、ilvD、pta、pur、pycA、tpi 7 个管家基因构建多位点序列分析分型方案,鉴定经随机扩增多态性DNA分型得到的42 株Bacillus cereus的序列类型。结果表明：42 株B. cereus分为7 个序列类型（sequence type,ST）,分别为ST-770（4.8%,2/42）、ST-1000（71.4%,30/42）、ST-1084（9.5%,4/42）、ST-1348（2.4%,1/42）、ST-1349（2.4%,1/42）、ST-1350（2.4%,1/42）和ST-1351（7.1%,3/42）;所有分离株被识别为205、142、23三个不同克隆谱系,2 个单态群（ST-770和ST-1351）和2 个独株（ST-1348和ST-1349）;发现4 个新的ST型和4 个新的等位基因,已上传至国际数据库得到新的序列号和等位基因号,分别为ST-1348、ST-1349、ST-1350、ST-1351和glp-253、glp-254、ilv-277、pyc-199。     

进出口食品中不同血清型沙门氏菌PFGE和 MLST分型比较研究

目的：通过PFGE和MLST两种方法对进出口食品中不同血清型沙门氏菌的分辨力和应用价值进行了比较研究。方法：采用PFGE和MLST两种方法对进出口食品中分离得到的鼠伤寒及肠炎两种血清型共30株沙门氏菌进行分型研究。结果：两种血清型共30株沙门氏菌经PFGE分型可得到23种型别,DI值为0.9563;鼠伤寒和肠炎两种血清型菌分别经PFGE分型,DI值分别为1.000和0.9048。MLST对30株沙门氏菌分型得到4种ST型,DI值为0.5793;鼠伤寒和肠炎两种血清型菌分别经MLST分型,DI值分别为0.2571和0.1333。结论：在分辨力方面,PFGE更适合于同一沙门氏菌血清型的分型,MLST适合于不同沙门氏菌血清型间的分型。MLST在替代、补充血清型分型方面有潜在的优势,PFGE在溯源研究方面似乎更胜一筹。     

Short communication: Molecular epidemiology of Streptococcus agalactiae differs between countries

Group B Streptococcus or Streptococcus agalactiae continue to be challenging for milk quality programs in countries with emerging dairy industries, such as Colombia, where high prevalence has been reported. Molecular typing of isolates is needed to understand the variability and epidemiology of this pathogen and to develop effective control and eradication programs. We characterized the molecular profile of Strep. agalactiae isolated from cows with subclinical mastitis in 21 Colombian dairy herds and measured diversity within and between herds using multilocus sequence typing. Isolates belonged to sequence type 248 [clonal complex (CC) 103; n = 30), ST1 (CC1; n = 6) or ST22 (CC22; n = 4)], whereas members of CC67/61, the dominant type in North America, were not detected. Presence of multiple clonally unrelated sequence type within a herd was common, which contrasts with the situation in European countries and suggests introduction from multiple sources. Our results demonstrate that conclusions from molecular epidemiological studies in 1 region cannot necessarily be extrapolated to other regions, and no single bovine-adapted CC of Strep. agalactiae exists in Colombia. Improvements in internal and external biosecurity will be needed to reduce Strep. agalactiae prevalence in Colombian dairy herds.     

A cohort study of the effect of Streptococcus agalactiae on milk yield and somatic cell count in Norwegian dairy cows

The primary objective of the present study was to estimate the effect of Streptococcus agalactiae intramammary infection on milk production and somatic cell count (SCC) in Norwegian dairy cows. A secondary objective was to assess differences in the effect of common Strep. agalactiae sequence types (ST) found in Norwegian dairy herds. We performed a cohort study combining registry data with sequence-type data from Strep. agalactiae isolates. Herds in which Strep. agalactiae had been detected in individual animals (bacteriological culture or quantitative PCR) between 2012 and 2015 were included. We accessed monthly test-day milk yield records for the entire period to compare milk yield and SCC between cows that were Strep. agalactiae positive and all other cows, within each herd. The study sample consisted of 150 herds, 15,757 cows, 30,850 lactations, and 204,126 test days. We evaluated the effects of Strep. agalactiae on test-day milk yield and SCC using mixed linear regression models, controlling for clustering by herd, cow, and lactation. Multilocus sequence typing of Strep. agalactiae was available for isolates from 86 herds. Additional models were fit to a subset of herds (n = 59) in which ST1, ST23, ST103, and ST196 had been found, to compare the effects of ST on milk production and SCC. In the period 3 to 2 mo before diagnosis, Strep. agalactiae-positive cows produced an average of 1.3 kg more DIM-adjusted milk/d than their negative herd mates. At the time of diagnosis, production was on average 0.13 kg less DIM-adjusted milk/d in Strep. agalactiae-positive cows than in negative cows; 2 to 3 mo after diagnosis, they produced 1.24 kg less DIM-adjusted milk/d than negative cows. Losses persisted for the rest of the investigated period. Cows with ST23, ST103, and ST196 followed a similar pattern as the overall analysis with respect to milk production, whereas ST1-affected cows produced similar amounts of milk before diagnosis as the negative cows. Cows with ST1 experienced the largest milk loss 1 to 2 mo after diagnosis but then recovered to some extent; for cows with ST103, the severe milk loss persisted for the rest of the investigation period. The cow-associated ST103 elicited a lower response in peak SCC compared with ST23, ST103, and ST196. The results indicate an effect of Strep. agalactiae on milk production and SCC. Production was lowest 2 to 3 mo after a positive sample. Peak SCC was reached the month before diagnosis, with notable differences between sequence types.     

Long-term persistence of specific genetic types of mastitis-causing Staphylococcus aureus on three dairies

Pulsed-field gel electrophoresis (PFGE) after SmaI digestion was used to investigate the persistence of specific genotypes of bovine mammary gland isolates of Staphylococcus aureus on 3 dairy herds. A total of 341 isolates of Staph. aureus were available from cows in 3 herds, collected over a period of 15 yr. Pulsed-field gel electrophoresis band patterns of Staph. aureus isolates were analyzed visually and with gel analysis and comparison software. Based on this analysis, isolates were classified by PFGE type. Persistence was determined as the time period from the first to the last isolation of a particular PFGE type of Staph. aureus within a herd. Specific types of mastitis-causing Staph. aureus persisted long-term on these dairies. For example, PFGE type 3 isolates persisted on farms A, B, and C for 15, 15, and 13 yr, respectively. Type 6 was found to persist for 13 yr on farm C. Despite the application of standard mastitis control practices, mastitis-causing Staph. aureus types appeared to persist long-term, as detected by PFGE, and were isolated coincident with herd problems of increased milk somatic cell counts and decreased milk production.     

我国婴儿配方粉来源的克罗诺杆菌脉冲场凝胶电泳分子分型和多位点序列分型研究

目的探讨我国婴儿配方粉污染克罗诺杆菌的主要分子分型特征,分析不同地区间菌株亲缘相关性,为我国克罗诺杆菌引起的疾病溯源提供技术支撑。方法采用脉冲场凝胶电泳(PFGE)和多位点序列分型(MLST)两种方法,对全国19个省/自治区/直辖市婴儿配方粉来源的49株克罗诺杆菌进行分型研究。结果 49株克罗诺杆菌PFGE分型得到38个带型,未发现有明显的优势分型和聚集现象。MLST共获得15个已知序列型(ST型)和2个新ST型,ST4、ST1和ST64为优势型别,分别占菌株总数的20.4%(10/49)、18.4%(9/49)、10.2%(5/49)。PFGE型和MLST型结果分析表明,具有相同PFGE型的菌株也有相同的MLST型,而相同MLST型却不一定具有较高的亲缘关系。结论我国婴儿配方粉来源的克罗诺杆菌图谱具有高多态性和离散性,据此预测我国婴儿配方粉污染克罗诺杆菌导致婴儿患病以散发为主,出现多人患病的暴发事件几率较低。     

出入境动物源性食品中单增李斯特菌的多位点序列分型分析

对出入境动物源性食品中分离的单增李斯特菌进行多位点序列分型（mutilocus sequence typing,MLST）分析,了解其序列型分布特点及不同菌株之间的亲缘关系。提取单增李斯特菌基因组DNA,选择其7 个管家基因进行聚合酶链式反应扩增并测序。将测序结果截成标准序列的长度后上传到MLST数据库进行比对分析,获得7 个管家基因的等位基因谱和序列分型编码,并将结果采用不加权算术平均组对（unweighted pair group method usingarithmetic averages,UPGMA）法进行聚类分析。89 株单增李斯特菌共获得51 个STs,其中26 个为新获得的STs（STnew1～STnew26）;数量最多的5 个STs为ST8（9.0%）,ST121（9.0%）、ST7（5.6%）、ST87（5.6%）及新发现的STnew3（7.8%）;其中ST456、ST34、ST343、ST19、ST517、ST201、ST98、ST330和ST73为在国内首次获得。采用UPGMA算法得到的进化树可将89 株菌株分为3 大类群,分类的结果与单增李斯特菌血清学家系分类结果一致。MLST结果对了解出入境动物源性食品中分离的单增李斯特菌的亲缘关系及流行病学溯源有重要意义。     

Genetic diversity of mastitis-associated Klebsiella pneumoniae in dairy cows

The objectives of this study were to determine the level of genetic diversity of Klebsiella pneumoniae isolated from clinical mastitis cases and to define genotypes most commonly associated with the disease. Individual quarter milk samples were collected from a single privately owned dairy herd over a 2-yr period and submitted to the Laboratory for Udder Health, Minnesota Veterinary Diagnostic Laboratory, University of Minnesota, for bacteriological culture. Eighty-four K. pneumoniae isolates were obtained and fingerprinted by repetitive DNA sequence PCR, 43 by pulsed-field gel electrophoresis (PFGE), and 29 by multilocus sequence typing (MLST). Significant genetic diversity was observed among the isolates regardless of the fingerprinting method used. Simpson's diversity index was 93.5, 96.1, and 97.0% when analyzed by repetitive DNA sequence PCR (n = 84), pulse field gel electrophoresis (n = 43), and MLST (n = 29), respectively. In some cases more than 1 genotype was obtained from a single milk sample originating from an individual quarter. The majority of infections were observed during the winter and accounted for 69.0% of K. pneumoniae mastitis cases. There was a negative correlation between a matrix of fingerprints similarity and a matrix of temporal distances. The MLST results revealed 5 new and novel allelic types, which have not been previously reported in the MLST database. Three isolates shared MLST types with human clinical isolates, raising the possibility that some K. pneumoniae isolates, of bovine origin, may be capable of causing disease in humans. There were 21 genotypes present within the herd, and there was no evidence for nonrandom distribution of genotypes uniquely associated with mastitis. We have shown, using 3 distinct genotyping methods, that K. pneumoniae isolated from clinical mastitis within a single dairy herd is caused by a genetically diverse population and that multiple genotypes can be isolated from a mastitic quarter. The data suggest that mastitis can be caused by a variety of K. pneumoniae genotypes. Diverse genotypes may have different levels of invasiveness and virulence and may originate from various sources within the dairy.     

Molecular characterization and clonal diversity of methicillin-susceptible Staphylococcus aureus in milk of cows with mastitis in Brazil

Mastitis is an important disease for the dairy industry worldwide, causing economic losses and reducing milk quality and production. Staphylococcus aureus is a worldwide agent of this intramammary infection, which also causes foodborne diseases. The objective of this study was to determine the frequency of methicillin-susceptible Staphylococcus aureus (MSSA) isolates in milk of mastitis cows in Brazil and to analyze the genetic lineages and the content of antimicrobial resistance genes and virulence factors among these isolates. Fifty-six MSSA isolates were recovered from 1,484 milk samples (positive for the California mastitis test) of 518 cows from 11 different farms in Brazil (representing 51% of total Staph. aureus obtained), and they were further characterized. Methicillin-susceptible Staphylococcus aureus were isolated from 3.7% of California mastitis test-positive tested milk samples and from 6.2% of tested mastitic cows. Methicillin-susceptible Staphylococcus aureus isolates were characterized by spa typing, agr typing, and multilocus sequence typing, and resistance and virulence traits were investigated by PCR. Seven spa types were identified among MSSA (% of isolates): t127 (44.6), t605 (37.5), t002, t1784, t2066 (1.8), and 2 new ones: t10856 (10.7) and t10852 (1.8). Five distinct sequence types (ST) were detected (% of isolates): ST1 (46.4), ST126 (37.5), ST133 (10.7), ST5 (3.6), and a novel ST registered as ST2493 (1.8). Resistances were detected for streptomycin, chloramphenicol, and tetracycline. One strain contained the chloramphenicol resistance gene (fexA; included within transposon Tn558) and 3 strains contained the tetracycline resistance gene [tet(K)]. Methicillin-susceptible Staphylococcus aureus strains were susceptible to most of the antibiotics studied and lacked the virulence genes of Panton-Valentine leukocidin (lukF/S-PV), toxic shock syndrome toxin 1 (tst), exfoliative toxin A (eta), and exfoliative toxin B (etb), as well as the genes of the immune evasion cluster. Methicillin-susceptible Staphylococcus aureus isolates were detected in a relatively low proportion of cows with mastitis (6.2%) and recovered isolates presented high diversity of genetic lineages, with CC1 and CC126 the predominant clonal complexes, and CC133 also being detected. Larger epidemiological studies with molecular characterization of isolates are required to deepen the knowledge on the circulating genetic lineages among the cow population with mastitis.     

Diagnostic accuracy of a standardized scheme for identification of Streptococcus uberis in quarter milk samples: A comparison between conventional bacteriological examination,modified Rambach agar medium culturing,and 16S rRNA gene sequencing

Bacteriological examination of milk samples is a prerequisite for pathogen-specific therapy and aids in limiting antimicrobial resistance. The aims of this study were to establish a standardized scheme for reliable Streptococcus uberis identification in routine diagnosis and to evaluate the accuracy of conventional tests and growing patterns of Strep. uberis on a selective medium (modified Rambach agar medium, MRAM) using 16S rRNA gene sequencing analysis as a reference method. We obtained isolates of presumptive Strep. uberis (n = 336) from quarter milk samples of dairy cows with intramammary infections and classified the isolates into 2 clusters using biochemical characterization. In cluster 1 (n = 280), cocci grew as non-hemolytic colonies, hydrolyzing esculin, carrying no Lancefield antigen (A/B/C/D/G) or Christie Atkins Munch-Petersen factor, and their growth was inhibited on an Enterococcus agar. Production of β-d-galactosidase on MRAM was shown by 257 of the cluster 1 isolates (91.79%), and 16S rRNA gene sequencing verified 271 (96.79%) of the isolates to be Strep. uberis. In 264 isolates (94.29%), MRAM agreed with the sequencing results. In cluster 2 (n = 56), isolates showed different characteristics: 37 (66.07%) were β-d-galactosidase-positive, and based on 16S sequencing results, 36 (64.29%) were identified correctly as Strep. uberis using biochemical methods. Identification success in this group differed significantly between routine diagnosis and MRAM application: MRAM agreed with sequencing results in 47 isolates (83.93%). To identify Strep. uberis and differentiate it from other lactic acid bacteria in routine diagnosis, we suggest using catalase reaction, hemolysis, esculin hydrolysis, and growth on enterococci agar. Isolates that show a typical biochemical profile can be identified satisfactorily with these tests. For Strep. uberis isolates with divergent patterns, application of MRAM as a follow-up test increased the diagnostic accuracy to 94.64%.     

Novel sequence types (STs) of Staphylococcus aureus isolates causing clinical and subclinical mastitis in flocks of sheep in the northeast of Brazil

Staphylococcus aureus is one of the most important infectious mastitis causative agents in small ruminants. In order to know the distribution of Staph. aureus strains associated with infectious mastitis in flocks of sheep in the northeast of Brazil and establish whether these clones are related to the strains distributed internationally, this study analysed the genetic diversity of Staph. aureus isolates from cases of clinical and subclinical mastitis in ewes by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). In this research, 135 ewes with mastitis from 31 sheep flocks distributed in 15 districts were examined. Staph. aureus was isolated from sheep milk in 9 (29%) out of 31 herds located in 47% of the districts surveyed. MLST analysis allowed the identification of four STs (ST750, ST1728, ST1729 and ST1730). The last three with their respective novel alleles (glp-220; pta-182 and yqil-180) were recently reported in the Staph. aureus MLST database (http://www.mlst.net). Each novel allele showed only a nucleotide different from those already described. The occurrence of CC133 (ST750 and ST1729) in this study is in agreement with other reports that only a few clones of Staph. aureus seem to be responsible for most cases of mastitis in dairy farms and that some of these clones may have broad geographic distribution. However, the prevalence of CC5 (ST1728 and ST1730)--an important group related to cases of colonization or infection in humans--differs from previous studies by its widespread occurrence and may suggest human contamination followed by selective pressures of the allelic diversifications presented for these STs.     

婴幼儿食品和配方奶粉中克罗诺杆菌污染调查及分子分型研究

目的了解温州市市售婴幼儿食品和配方奶粉中克罗诺杆菌污染情况及分子分型特征,分析不同菌株间亲缘相关性。方法参照GB 4789.40—2010《食品安全国家标准食品微生物学检验阪崎肠杆菌检验》方法在食品中分离鉴定克罗诺杆菌。采用脉冲场凝胶电泳(PFGE)和多位点序列分型(MLST)方法对克罗诺杆菌进行分型研究。结果 2013—2017年共采集737份婴幼儿食品和配方奶粉,克罗诺杆菌总检出率为6.1%(45/737)。其中婴幼儿谷类辅助食品检出率最高(23.4%,37/158)。45株克罗诺杆菌属于阪崎克罗诺杆菌(C.sakazakii)、丙二酸盐克罗诺杆菌(C.malonaticus)和莫金斯克罗诺杆菌(C.muytjensii)。其中阪崎克罗诺杆菌最多(73.3%,33/45),其次为丙二酸盐克罗诺杆菌(24.4%,11/45)。MLST共分为25个ST型,其中ST64、ST1、ST40、ST4和ST7为优势型别。45株克罗诺杆菌PFGE分型得到38个带型,未发现有明显的优势分型和聚集现象。PFGE型和MLST型结果分析表明,绝大数具有相同PFGE型的菌株也有相同的MLST型,而相同MLST型却不一定具有较高的亲缘关系。结论温州市市售婴幼儿食品和配方奶粉中克罗诺杆菌的污染状况较轻,其中婴幼儿谷类辅助食品检出率最高。食品来源的克罗诺杆菌图谱具有高多态性和离散性。     

Host adaptation of bovine Staphylococcus aureus seems associated with bacteriological cure after lactational antimicrobial treatment

Staphylococcus aureus causes a wide range of diseases in multiple species. Some sequence types (ST) are observed in a variety of hosts, whereas other strains are mainly associated with bovine mastitis, suggesting host adaptation. We propose that host adaptation of Staph. aureus may influence bacteriological cure of bovine subclinical mastitis after antimicrobial treatment. To test this hypothesis, multilocus sequence typing was performed on Staph. aureus isolates from 60 treated and 79 untreated control quarters that were obtained from well-defined cohorts of dairy cows from a recently conducted randomized field trial on early treatment of subclinical mastitis. Bovine-associated ST were distinguished from non-bovine-associated ST based on the literature and public databases. The association between host adaptation and bacteriological cure was investigated using population-averaged logistic regression models. Thirteen ST were identified, with approximately 80% of isolates belonging to bovine-associated ST. The odds for cure were around 2.5 times as high for non-bovine-associated ST as for bovine ST in treated quarters, whereas no difference in spontaneous cure was observed in untreated control quarters. In addition, host adaptation was related to known predictors of cure, such as penicillin susceptibility and somatic cell count. All isolates belonging to non-bovine-associated ST were resistant to penicillin, whereas the majority of isolates belonging to bovine-associated ST were penicillin susceptible. Penicillin-resistant bovine-associated strains were associated with high somatic cell counts compared with other strains. The correlation between penicillin resistance, cell counts, and host adaptation may affect the association between host adaptation and cure. For diagnostic purposes, a simple and fast alternative to multilocus sequence typing of Staph. aureus to determine host adaptation may be valuable.