2016‒2017年辽宁省沙门菌耐药菌谱与分子分型

摘要：目的 分析2016?2017年辽宁省沙门菌分离株的耐药特性与脉冲场凝胶电泳（PFGE）分子分型特征,为沙门菌引起的食源性疾病暴发、防控及抗生素使用提供参考数据。方法 对分离的54株沙门菌进行血清分型和药物敏感试验。根据PulseNet沙门菌标准PFGE分型技术,选取全部菌株进行PFGE分子分型分析,应用BioNumerics软件对菌株条带进行分析,确定菌株间的特征及相关性。结果 54株沙门菌血清型居首位的是肠炎沙门菌,占46.30％;其次是鼠伤寒沙门菌,占24.07％;共分为10个血清型。对13种抗生素的耐药分析显示多重耐药菌株为36株,占66.7％,其中耐3～5种的13株（24.1%）,耐6～8种的13株（24.1%）,耐9～11种的10株（18.5%）。54株沙门菌经聚类分析获得36种带型,相似度区间为49.7%～100.0％。结论 辽宁省沙门菌分离株多重耐药状况比较严重,相同血清型其PFGE带型相似度相对较高,同时具有较显著的优势带型特点;而且发现同一PFGE型菌株的耐药谱相对比较接近。     

Plasmid characteristics of Salmonella strains of various origins

Salmonella antibiotic-resistant strains, isolated from patients with hospital infections and from various environmental objects, showed lower virulence than antibiotic-sensitive strains in experiments on mice infected by intraperitoneal and enteral routes. Salmonella strains, sensitive to antimicrobial preparations, contained 1-2 plasmids, while those with multiple drug resistance contained 3-10 plasmids varying in their molecular weight. All these strains, with the exception of one laboratory strain, carried a plasmid with a molecular weight of about 60 Md. A decrease in the virulence of Salmonella strains, carrying R-plasmid, with respect to mice, their natural host, in experimental infection by the above-mentioned routes was probably unrelated to the loss of this plasmid. 80% of Salmonella strains with multiple resistance to antibiotics yielded positive results in the keratoconjunctival and conjunctival tests as compared with 42% of sensitive strains. These data suggest that Salmonella strains, carrying R-plasmid, retained pronounced capacity for local colonization.     

辽宁食源性沙门菌血清型、 耐药谱及PFGE分型特征

目的分析辽宁食源性沙门菌血清型、耐药谱及脉冲场凝胶电泳(PFGE)型别,探讨辽宁沙门菌污染的同源性,为食源性疾病溯源和预警提供基础。方法对辽宁省2015年食品中、食源性疾病中分离的41株沙门菌进行血清学分型、耐药试验、PFGE分子分型,采用Bio Numerics version 6.6软件分析,比较同源性。结果 41株菌分为15个血清型,居前三位的是15株肠炎沙门菌、5株德尔卑沙门菌、5株姆班达卡沙门菌(辽宁省内少见血清型);对41株菌进行15种抗生素的耐药试验,对单一一种抗生素的耐药率为100.0%,其中红霉素97.6%,萘啶酸61.0%,氨苄西林53.7%;41株菌共分为18种PFGE带型,带型分布分散,只有两种优势,一种带型包含20株菌,有14株肠炎沙门菌,6株其他沙门菌,相似度为92.7%~100%;另一种包含5株菌,4株姆班达卡沙门菌,1株鼠伤寒沙门菌,相似度为96.6%~100.0%。结论辽宁省食源性沙门菌的血清型以肠炎沙门菌为主,生肉制品是其主要污染来源;血清型与PFGE图谱带型分布广泛,相同血清型沙门菌的PFGE带型聚集成簇、菌株具有高度同源性;相同PFGE型别的菌株耐药谱一致或相似;沙门菌的耐药情况较严重。     

Antibiotic resistance of Salmonella strains isolated from natural polluted waters

Resistance to 14 antibiotics was tested in 270 Salmonella strains isolated from different aquatic environments. All the strains were sensitive to nalidixic acid (30 micrograms) and cephalothin (30 micrograms) but more than 90% were resistant to tetracycline (30 micrograms). The percentage of strains resistant to other antimicrobial substances depended on the antibiotic and on the isolation source. Twenty-four resistance patterns were recorded in strains isolated from three environmental sources. The only multi-resistance detected in the three ecosystems was that of sulphadiazine and tetracycline (about 20%). The serotypes most frequently detected with multi-resistance to different antibiotics were Salmonella typhimurium and Salm. blockley.     

Transmissive drug resistance and virulence of "epidemic" strains of S. typhimurium]

A total of 59 "epidemic" strains of S. typhimurium and 52 cultures isolated from the cases with sporadic diseases were studied with respect to the nature of their resistance to 10 antibiotics and virulence for albino mice under conditions of subcutaneous and oral inoculation. The virulence of the cultures isolated from the cases with sporadic diseases was higher in the strains sensitive to antibiotics. The "epidemic" strains were characterized by the lowest virulence for the mice and resistance to 8-10 antibiotics simultaneously with the transmissive nature of resistance to 1-5 drugs. The transmissive nature of resistance to antibiotics and its spectrum may serve an additional epidemiological marker of the strains.     

Antimicrobial resistance of Salmonella spp. from pigs at slaughter in Spain in 1993 and 2001

AIM: To compare the incidence of antimicrobial resistance among Salmonella serotypes isolated in a pig slaughterhouse in Zaragoza (Spain) during 1993 and 2001. METHODS AND RESULTS: A total of 168 isolates representing 10 serotypes were examined by disc diffusion method using 17 antibiotics. Data showed that the majority of the strains were resistant to streptomycin (97%), sulfadiazine (93.4%) and tetracycline (83.3%). A large proportion of the collection was multidrug resistant (MDR, resistance to four or more antibiotics) with a greater incidence in 2001. The findings imply an increasing incidence of MDR amongst S. Typhimurium, and all S. Typhimurium-definitive phage type (DT) 104 isolates were resistant to ampicillin, chloramphenicol, streptomycin, sulphonamide and tetracycline (R-ACSSuT). This resistance phenotype had spread among other phage and serotypes. Salmonella Ohio was also a MDR serotype and this is not a serotype normally associated with drug resistance. CONCLUSIONS: A large proportion of the strains were MDR and this showed that pork products could be a potential vehicle of MDR Salmonella food-borne infections. SIGNIFICANCE AND IMPACT OF THE STUDY: The findings may have significant public health consequences and could contribute to the development of useful practices aimed at limiting the transmission of MDR Salmonella serotypes through the food chain.     

Antibiotic resistance of Salmonella strains isolated from natural polluted waters

Resistance to 14 antibiotics was tested in 270 Salmonella strains isolated from different aquatic environments. All the strains were sensitive to nalidixic acid (30 μg) and cephalothin (30 μg) but more than 90% were resistant to tetracycline (30 μg). The percentage of strains resistant to other antimicrobial substances depended on the antibiotic and on the isolation source. Twenty-four resistance patterns were recorded in strains isolated from three environmental sources. The only multi-resistance detected in the three ecosystems was that of sulphadiazine and tetracycline (about 20%). The serotypes most frequently detected with multi-resistance to different antibiotics were Salmonella typhimurium and Salm. blockley.     

Parallel evolution of multidrug-resistance in Salmonella enterica isolated from swine

The increase in frequency of Salmonella enterica resistant to antibiotics in food-producing animals is of great concern to public health. Determining the rate at which different resistance phenotypes are generated and maintained in the environment is thus of great importance. The distribution and evolution of antibiotic resistance and multidrug-resistance in 362 Salmonella stains as part of a cross-sectional study of the Canadian swine industry were investigated. The susceptibility of all isolates to 12 antimicrobial agents was tested and the statistical and phylogenetic distribution of resistance among strains characterized via multilocus sequence typing was studied to test the origin of multidrug-resistance in Salmonella. More than 25% of all isolates were multidrug-resistant, with predominance in serotype Typhimurium, a serotype of vital importance to public health. The strong associations between resistance phenotypes, which differ among serotypes and which is supported by the significant genetic distance between serotypes, was indicative of the independent acquisition of multidrug-resistance in at least two different serotypes, i.e. Typhimurium and Derby. The independent origin of multidrug-resistance in Salmonella indicates that strong selective pressures are present in the environment of the bacteria and that statistical and phylogenetic studies of antibiotic resistance are an essential part in the understanding and the control of the epidemic.     

Escherichia coli strains isolated from surface waters. Distribution by resistance to antibiotics and R-plasmid transfer

Samples were taken from surface waters in Csongrád county in the months March through December, 1980. Escherichia coli isolates selected at random were tested for resistance to 5 antibiotics and for R-plasmid carriership. Of the strains isolated from Tisza river at each of 8 sampling sites, 50-60% were sensitive to all the 5 antibiotics. The percentage of sensitivity was much lower if only the strains isolated during the summer months, when the water level was high, or those isolated from affluents and backwaters were taken into account. The frequency of resistance was the highest for tetracycline, followed in order by ampicillin, streptomycin, kanamycin and chloramphenicol. R-plasmid was carried by 43% of the resistant isolates tested, mainly by multiresistant ones.     

Comparison of virulence factors and R plasmids of Salmonella spp. isolated from healthy and ill swine.

The antibiotic resistance and virulence profiles of Salmonella spp. isolated from healthy (group 1) and ill (group 2) swine were compared. Parameters studied included colicin and siderophore production; mannose-sensitive hemagglutination of erythrocytes; resistance to the lethal effect of serum complement; resistance to antibiotics; and the transmissibility of these characteristics to recipient organisms. Group 1 (19 isolates) had 14 serotypes, and group 2 (20 isolates) had 2 serotypes. Isolates from group 2 were resistant to more antibiotics and had a greater ability to hemagglutinate erythrocytes and transfer R plasmids to recipient organisms, but a lesser ability to produce siderophore than group 1. All 39 isolates resisted the lethal effects of serum complement. Colicin was produced by 1 of 19 from group 1 and 0 of 20 from group 2. A donor Escherichia coli isolated from a pig with enteritis transferred R plasmids to 62% of group 1 and 0% of group 2 Salmonella spp. when they were used as recipient organisms. A transconjugant from the mating of donor E. coli to a group 1 Salmonella spp. was further able to pass an R plasmid to recipient E. coli and salmonellae. Plasmid isolation from group 1 yielded 1 of 19 strains with a 56-megadalton plasmid, while 20 of 20 strains from group 2 contained three to five plasmids from 2.4 to 60 megadaltons in size.     

Comparison of virulence factors and R plasmids of Salmonella spp. isolated from healthy and ill swine

The antibiotic resistance and virulence profiles of Salmonella spp. isolated from healthy (group 1) and ill (group 2) swine were compared. Parameters studied included colicin and siderophore production; mannose-sensitive hemagglutination of erythrocytes; resistance to the lethal effect of serum complement; resistance to antibiotics; and the transmissibility of these characteristics to recipient organisms. Group 1 (19 isolates) had 14 serotypes, and group 2 (20 isolates) had 2 serotypes. Isolates from group 2 were resistant to more antibiotics and had a greater ability to hemagglutinate erythrocytes and transfer R plasmids to recipient organisms, but a lesser ability to produce siderophore than group 1. All 39 isolates resisted the lethal effects of serum complement. Colicin was produced by 1 of 19 from group 1 and 0 of 20 from group 2. A donor Escherichia coli isolated from a pig with enteritis transferred R plasmids to 62% of group 1 and 0% of group 2 Salmonella spp. when they were used as recipient organisms. A transconjugant from the mating of donor E. coli to a group 1 Salmonella spp. was further able to pass an R plasmid to recipient E. coli and salmonellae. Plasmid isolation from group 1 yielded 1 of 19 strains with a 56-megadalton plasmid, while 20 of 20 strains from group 2 contained three to five plasmids from 2.4 to 60 megadaltons in size.     

Prevalence of Salmonella enterica serovars and genovars from chicken carcasses in slaughterhouses in Spain

AIMS: To determine the prevalence of Salmonella enterica serovars in chicken carcasses in slaughterhouses in Spain and to examine genotypic relations among these serovars. METHODS AND RESULTS: A total of 336 chicken carcasses were collected from six slaughterhouses in Northwestern Spain. Salmonellae were isolated (ISO-6579-1993), serotyped, phage-typed, ribotyped and antibiotyped against 20 antibiotics. Salmonella strains were detected in 60 (17.9%) carcasses. Isolates belonged to nine different serotypes, with Salm. Enteritidis being the most common. Three strains (5%) were resistant to one antibiotic and 24 (40%) were multi-resistant (to more than one antibiotic). The most frequently encountered resistances were to sulphamides, fluoroquinolones and tetracycline. Ribotyping was able to differentiate isolates of the same serotype and phage type. CONCLUSIONS: The Salmonella serotypes and phage types detected are among those most frequently associated with human diseases in Spain. The large percentage of antimicrobial resistant strains is a matter for concern. A high genetic relationship between strains from different slaughterhouses was found. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides detailed information about Salmonella isolates from poultry in Spain. It emphasizes the importance of controlling this pathogen in poultry products, and suggests the need for more prudent use of antibiotics.     

Antibiotic resistance of Salmonella during 1982 and 1983

The antibiotics susceptibility of 480 Salmonella collected in 1982 and 1983, in the National Center of Salmonella of Pasteur Institute of Tunis was tested. High levels of resistance were found. Nalidix acid, colistin and gentamicin were the most active. Resistance and multiple resistance was most frequently found in strains of Salmonella wien and Salmonella saint paul.     

Multidrug resistance to antibacterial drugs of Salmonella enterica subsp. enterica strains isolated in Poland in the 1998-1999 period]

A total of 510 Salmonella enterica subsp. enterica strains representing 56 serotypes, isolated from human stool specimens during 1998-2000 in sanitary-epidemiological units in Poland were tested for their susceptibility by a standard disk diffusion method for: ampicillin, cefotaxime, chloramphenicol, tetracycline, streptomycin, gentamicin, kanamycin, nalidixic acid, ciprofloxacin, furazolidone, cotrimoxazole, sulfonamides and trimethoprim. For 201 of the investigated strains, belonging to 5 most common isolated serotypes (S. Enteritidis, S. Typhimurium, S. Hadar, S. Infantis and S. Virchow) the minimal inhibitory concentrations (MICs) for the aforementioned antibiotics, as well as for amoxicillin with clavulanian were determined. Selected strains were screened for production extended spectrum b-lactamases (ESBLs). It was observed that 42.9% of Salmonella enterica subsp. enterica strains were resistant to 2 or more antibiotics, with the highest prevalence of MDR strains among serotypes Typhimurium, Hadar and Virchow. Resistance to ampicillin, streptomycin, tetracycline, nalidixic acid, furazolidone and sulphonamides was observed most frequently. Over 93% of S. Virchow strains were resistant to furazolidone. No strains resistant to ciprofloxacin were detected according to the NCCLS guidelines, but 31.3% of isolates exhibiting reduced ciprofloxacin susceptibility (MICs ranging between 0.125 and 0.5 mg/l). Two strains S. Mbandaka and Salmonella group D (variant motility--) were resistant to cefotaxime and probably produced ESBL.     

Incidence of antibiotic resistance in Salmonella typhimurium strains isolated in different epidemic situations

A total of 1800 S. typhimurium strains isolated in different regions of the USSR in 1968-1979 were studied. Of these strains, 68.6% were resistant to 6-8 antibiotics, 7.8% were resistant to 2-5 antibiotics, and 23.6% proved to be resistant to all antibiotics. The number of multiresistant strains sharply increased during the last 10 years. The strains isolated from different sources and in different epidemic situations considerably differed in their sensitivity to the action of antibiotics. The infective agents isolated from the hospital foci of salmonellosis were found to possess the maximum multiresistance. The study of the genetic nature of multiresistance showed that the multiresistance strains had a conjugative R-plasmid with resistance determinants CmTc, type Fin+, incompatibility group F1me. The phage typing of the strains with phages from the collection of the Tbilisi Research Institute for Vaccines and Sera revealed that these strains belonged to phagotype 2. The problems of the relationship between the biological properties of causative agents and the character of the epidemic process of the diseases caused by these agents are discussed.     

Resistance of Salmonellae Isolated in 1959 and 1960 to Tetracyclines and Chloramphenicol

The resistance of cultures of Salmonella typhimurium to tetracyclines and chloramphenicol has been examined periodically. Although none of 200 cultures isolated prior to 1948 was resistant to the antibiotics, 5% of 100 cultures from man and 9% of 100 cultures from fowls which were isolated in 1956 and 1957 were resistant to tetracyclines. Among 158 cultures isolated from man and 100 cultures isolated from fowls in 1959 and 1960, 13.9 and 29%, respectively, were resistant to tetracyclines. In the last series, cultures resistant to chloramphenicol were found for the first time. Among 150 cultures of other Salmonella serotypes from man and 137 similar cultures isolated from fowls in 1959 and 1960, 5.3 and 8%, respectively, were found resistant to tetracyclines. There is no obvious explanation for the higher percentage of resistant strains occurring in S. typhimurium than in other serotypes.     

Aquarium pets as a source of antibiotic-resistant salmonellae.

Thirteen serotypes of Salmonella isolated from imported ornamental aquarium frogs, snails, and their waters were shown to be multi-drug-resistant. Among the resistances exhibited were resistance to gentamicin, sulfamethoxazole-trimethoprim, cephalothin, and nalidixic acid. Frog isolates displayed eight different patterns and snails isolates had two different resistance patterns. The most common serotype, Salmonella typhimurium, was resistant to 18 antibacterials while other salmonellae were resistant to 9 to 16 antibacterials. Resistances in S. typhimurium and S. bovis-morbificans were conjugative and a number of R plasmids participated in the resistance. The plasmid-mediated resistance in S. typhimurium was stable and the levels of resistance conferred were markedly higher than in the other salmonellae tested. Resistance of other serotypes was non-conjugative and resistance to the beta-lactam antibiotics was unstable.     

鸡蛋生产链中沙门氏菌对抗生素及消毒剂的耐药性研究

为研究鸡蛋生产链中沙门氏菌的污染情况及抗生素、消毒剂耐药情况,本文鉴定了鸡蛋生产链中分离得到的111株沙门氏菌(Salmonella)血清型,并测定了抗生素和消毒剂对沙门氏菌的最小抑菌浓度(Minimum inhibitory concentrations, MICs),检测了其对抗生素和消毒剂的耐药基因的表达情况。研究结果表明,沙门氏菌对甲氧苄啶(Trimethoprim, TMP)耐药率最高(N=100,P=90.09%),对阿莫西林/克拉维酸(Amoxicillin and clavulanate, AMC)、头孢噻呋钠(Sodium ceftiofur, CFS)、庆大霉素(Gentamicin, CN)敏感。沙门氏菌共产生6种不同的耐药谱型,TMP是最主要的耐药谱型(N=36,P=32.43%),52.25%的菌株(N=58)具有多重耐药性。苯扎氯铵(Benzalkonium chloride, BC)与氯化十六烷基吡啶(Cetylpyridinium chloride, CPC)对沙门氏菌的MIC的范围分别为：8~128 μg/mL、8~256 μg/mL。相对于质控菌株Escherichia coli ATCC10536,101株沙门氏菌对BC和CPC同时具有较高的耐药性(P=90.99%),109株沙门氏菌对抗生素和消毒剂具有共同耐药性(P=98.20%)。抗生素耐药基因检出率最高为blaTEM(N=49, P=44.14%),未检测出qnrA、qnrB、qepA基因,仅检测出qacEΔ1消毒剂耐药基因(N=63, P=56.76%)。抗生素耐药基因sul1和消毒剂耐药基因qacEΔ1具有显著相关性(P<0.01)。S. Derby对TMP、土霉素(Oxytetracycline, OTC)、阿莫西林(Amoxicillin, AML)、环丙沙星(Ciprofloxacin, CIP)同时表现较高的耐药性,S. Derby检出了11种抗生素耐药基因,消毒剂耐药基因qacEΔ1的检出率为81.25%(N=52)。鸡场中养殖内环境沙门氏菌对抗生素和消毒剂的耐药率以及耐药基因检出率均高于养殖外环境,鸡蛋包装、储存及销售等环节中沙门氏菌耐药率及耐药基因检出率均较高。由此可见,鸡蛋生产链中沙门氏菌对抗生素、消毒剂耐药性较严重,且存在共同耐药的现象。因此,需要进一步规范防控鸡场中沙门氏菌,规范抗生素和消毒剂的使用以及加强鸡蛋生产链条中卫生安全的监管。     

临床和非临床分离沙门菌的菌型分布及对抗生素的耐药性研究

目的：探讨沙门菌在腹泻病人粪便、外环境污水和部分动物标本中的菌型分布及其对抗生素的耐药性。方法：采用血清学方法对沙门菌进行分群和分型,采用K—B法进行抗生素耐药性测定。结果：410份临床腹泻病人标本和514份非临床标本共检出沙门菌93株,检出率为10．1％,其菌型分布为：腹泻病人粪便中检出的均为B群鼠伤寒沙门菌(6／93),非临床标本中的菌型主要为C2群纽波特沙门菌(72／93),还有E1群伦敦沙门菌(7／93)、B群鼠伤寒沙门菌(2／93)和未定型(6／93)。87株沙门菌对青霉素的耐药率最高(96．5％),其次为SMZ TMP(73．6％),未发现对新霉素、链霉素、四环素和阿米卡星的耐药株。结论：实验结果可为研究我省沙门菌的菌型分布及其对抗生素的耐药性提供参考;临床上应加强对鼠伤寒沙门菌引起腹泻的监测。     

R plasmid distribution in S. typhi isolated on the territory of the left coast of the Ukraine in 1970-1982

A total of 373 strains of S. typhi isolated in 1970-1982 were tested with respect to their sensitivity to 9 antibiotics active against gram-negative bacteria. It was shown that about 1/3 of the isolates were resistant to 1-3 antibiotics. Among the resistant isolates the number of strains resistant to 4-5 and more antibiotics amounted to 12.5 per cent. The plasmid nature of the antibiotic resistance in the isolated strains of S. typhi was shown. Transmissive R plasmids were detected in 13 per cent of the strains studied. Within the last 5 years there was an increase in the proportion of strains with transmissive R plasmids in patients with sporadic typhoid fever or especially in groups of patients with the disease.