1993-2000年霍乱弧菌耐药整合子的分布状况

目的检测Ⅰ类、Ⅱ类和Ⅳ类整合子及Ⅰ类整合子携带的耐药基因盒在霍乱弧菌临床菌株中的分布，分析整合子对细菌耐药性的影响。方法纸片扩散法行药物敏感试验，PCR法检测50株临床菌株Ⅰ类、Ⅱ类和Ⅳ类整合酶基因（intⅠ）阳性菌株，并对intⅠ1阳性菌株整合的耐药基因行序列分析。结果全部菌株至少对一种抗生素耐药。3株Ⅰ类整合酶基因阳性菌株的耐药基因盒PCR扩增得到1009bp的产物。序列分析表明，1009bp序列与已知的aadA 1c 100％同源，为对壮观霉素、链霉素产生耐药的基因盒；未检测到Ⅱ类整合子阳性菌株；50株临床菌株均含有第Ⅳ类整合酶基因，2株菌株第Ⅳ整合酶基因扩增得到2200bp产物，序列分析表明，在第Ⅳ类整合酶基因序列中含有一个插入片段IS1359转座酶基因。结论第Ⅰ类整合子存在于孟加拉国的临床致病弧菌中，第Ⅳ类整合子在霍乱弧菌临床菌株中分布广泛，整合子在细菌耐药中发挥作用。     

医院污水中大肠埃希菌的耐药谱及Ⅰ类整合子分析

目的调查医院污水中多重耐药菌污染情况及其耐药基因和Ⅰ类整合子携带情况。方法采集河南省郑州大学第一附属医院污水,使用16S rRNA测序和微量生化反应管鉴定细菌种属;采用PCR扩增耐药基因和Ⅰ类整合子基因盒,计算其检出率。结果根据16S测序和生化反应共分离鉴定出23株大肠埃希菌。药敏试验显示23株大肠埃希菌对四环素的耐药率为62.5%(15/23),复方新诺明为56.5%(13/23),氨苄西林为56.5%(13/23)。所有菌株均对美罗培南和亚胺培南敏感。大肠埃希菌多重耐药率为56.52%(13/23)。PCR检测耐药基因携带率aadA1基因为100%,tetA基因为95.8%,aac(6')-Ib-cr基因为79.2%,CTX基因为70.8%,tetB基因为63.5%,Qnrs基因为58.3%,SHV基因为54.2%,qepA基因为41.7%,Sul基因为35%。所有菌株均不携带KPC基因、IMP基因和NDM1基因。有21株细菌检测到Ⅰ类整合子,其中11株细菌的整合子携带空基因盒。整合子携带的大多数基因盒编码DfrA和aadA基因,有1株细菌的基因盒编码OXA-1基因。结论医院污水中的大肠埃希菌存在多药耐药现象且Ⅰ类整合子携带率高,应引起高度重视。     

整合子及相关基因盒在临床分离铜绿假单胞菌中的分布

目的检测Ⅰ类、Ⅱ类和Ⅲ类整合子及Ⅰ类整合子相关基因盒在铜绿假单胞菌临床分离株中的分布,分析整合子对细菌耐药性的影响。方法用纸片法对62株临床分离铜绿假单胞菌进行药敏试验;应用多重PCR法检测62株铜绿假单胞菌Ⅰ类、Ⅱ类和Ⅲ类整合子;对Ⅰ类整合子阳性菌进行整合子相关基因盒检测。结果62株菌中有40株(64.5%)含有Ⅰ类整合子,1株(1.6%)含有Ⅱ类整合子;没有检测到Ⅲ类整合子阳性菌。在Ⅰ类整合子阳性菌中,有26株携带Ⅰ类整合子相关基因盒(65.0%);分离自同一科室的部分菌株携带大小相同的基因盒;Ⅰ类整合子阳性菌株的耐药率高于整合子阴性的菌株。结论Ⅰ类整合子及整合子相关基因盒在铜绿假单胞菌临床菌株中分布广泛,整合子在细菌耐药中发挥作用。     

临床痢疾志贺菌整合子检测及耐药基因盒序列分析

目的 检测印度东部1988、1995和2002年部分临床分离痢疾志贺菌中细菌耐药关系密切的1、2、3类整合酶基因及整合子携带的耐药基因盒的分布,分析整合子系统对志贺菌耐药的影响.方法 纸片扩散法检测实验菌株对药物的敏感性;应用PCR方法对16株临床耐药菌株进行1、2、3类整合酶基因(intI)筛选,对阳性样本可变区基因盒序列进行鉴定分析.结果 所有16株菌均耐4种及4种以上药物,包括β-内酰胺类、氨基糖苷类、四环素类、磺胺类、氯霉素类和喹诺酮类.13株菌检出1类整合酶基因,全部菌株含2类整合酶基因,即发现同时存在两种整合子结构菌株,未检测到3类整合酶基因.1类整合酶插入基因盒以blaara30-aadAl基因家族为主,分别对β-内酰胺类抗生素、链霉素、壮观霉素耐药;2类整合酶插人基因盒以dfrAl-satl组合为主,分别对甲氧苄氨嘧啶、链丝菌素耐药,同时在4株菌中发现dfrAl-satl-aadAl基因盒组合.结论 2类整合子普遍存在于临床志贺菌中.整合子与志贺菌的多重耐药具有密切相关性.     

耐药志贺菌1类整合子的研究

目的分析我院志贺菌的流行菌株类型和耐药特点,研究志贺菌1类整合子的检出率、与耐药的相关性及其携带的耐药基因盒。方法从本院肠道门诊腹泻患者的粪便标本分离出70株志贺菌,以K-B琼脂法测定药敏情况;PCR扩增1类整合子可变区并进行序列测定和基因盒分析。结果70株志贺菌对四环素、复方新诺明、氨苄西林、利福平、萘啶酸的耐药率均>60%;对喹诺酮类常用药诺氟沙星耐药率较低,为12.8%;对头孢菌素类的耐药率≤10%。多重耐药率较高,达到77.1%。16株(22.9%)志贺菌中检出1类整合子。其中8株检出大小约为1 900 bp的1类整合子,测序显示含有编码对甲氧苄啶耐药的基因盒dhfrXⅡ和对氨基糖苷类耐药的基因盒aadA2,另8株检出大小约为1 000 bp的1类整合子,测序显示含有编码对氨基糖苷类耐药的基因盒aadA2。结论本院志贺菌流行株主要是福氏和宋内氏志贺杆菌,多重耐药率较高。志贺菌中存在1类整合子,与志贺菌的耐药具有相关性     

产超广谱β内酰胺酶临床分离株可转移多重耐药分子机制的研究

目的 研究产超广谱β内酰胺酶(ESBLs)临床分离株可转移多重耐药性的分子机制。方法 采用E-试验条进行药敏检测，电转化试验筛选、分离耐药质粒，聚合酶链反应(PCR)扩增Ⅰ型整合子基因盒插入序列及其分子克隆和序列分析。结果 9个产ESBLs耐药质粒中有8个检测出了插入序列，其中7个携带了1～2种抗药性基因盒。包括氨基糖苷类钝化酶aacA4、aadA2和aadA5；甲氧苄啶钝化酶dhfrA12和dfrA17；利福平钝化酶arr-3；氯霉素外排蛋白cmlA6。基因盒功能与转化子耐药表型一致。结论 质粒定位和整合子介导的抗药性基因盒可能是导致ESBLs产酶株多重耐药性产生和(或)播散的重要原因。     

肉鸡源多重耐药空肠、结肠弯曲菌的耐药分子特征

目的了解肉鸡源空肠和结肠弯曲菌的耐药谱特征,检测多重耐药菌株Ⅰ类整合子/基因盒、gyrA基因突变位点、tetO基因、23S rRNA突变位点的分子特征。方法利用PCR检测弯曲菌Ⅰ类整合子/基因盒的存在情况;利用MAMA PCR技术检测弯曲菌gyrA基因第257位碱基的突变情况;针对弯曲菌23S rRNA的V区2075突变位点检测突变菌株。结果多重耐药菌株占分离株的94.5%。146株多重耐药空肠和结肠弯曲菌中Ⅰ类整合子检出率为98.6%,有78株菌株检出3种基因盒,1 000 bp+750 bp+500 bp+250 bp为主要谱型,所占比例为92.3%;有131株在gyrA喹诺酮类耐药决定区发生突变,突变率为92.9%。127株四环素耐药弯曲菌tetO基因的检出率为95.3%。81株红霉素耐药菌株中,23S rRNA的V区2075处突变发生率为96.3%。结论空肠和结肠弯曲菌分离株携带aadA2耐药基因盒,与氨基糖苷类药物的耐药性相关;gyrA基因突变、tetO基因的携带以及23S rRNA突变,与弯曲菌对喹诺酮、四环素和大环内酯类耐药密切相关。     

耐三代头孢菌素阴沟肠杆菌临床株中CTX-M型ESBLs与Ⅰ类整合子的相关性

目的研究CTX-M型超广谱β内酰胺酶（ESBLs）及Ⅰ类整合子在耐三代头孢菌素阴沟肠杆菌中的分布,进一步探讨Ⅰ类整合子与CTX-MM型ESBLs的关系.方法运用K-B法检测阴沟肠杆菌临床株的耐药表型,双纸片协同试验（DDST）初筛产ESBLs的菌株,聚合酶链反应（PCR）扩增确证产CTX-M型ESBLs和含有Ⅰ类整合子的临床菌株,套式PCR及序列测定寻找携带CTX-M型ESBLs基因盒的Ⅰ类整合子.结果37株耐药菌中,21株菌产生CTX-M型ESBLs;20株菌含有Ⅰ类整合子;在13株同时产生Ⅰ类整合子和CTX-M型ESBLs的临床株中,3株菌CH4、CH11和Q1的CTX-M型ESBLs基因盒分布在Ⅰ类整合子上.结论Ⅰ类整合子的存在增加了ESBLs在临床株中水平播散的危险,是造成多重耐药株在院内暴发流行的重要原因.     

鲍曼不动杆菌Ⅰ类整合子和ISCR1的检测及其结构分析

目的了解鲍曼不动杆菌中Ⅰ类整合子与ISCR1分布情况及其所携带耐药基因的种类,从而分析其结构及其与耐药性的关系。方法采用煮沸法提取鲍曼不动杆菌基因组DNA,设计引物进行PCR反应,扩增产物用1.2%琼脂糖凝胶电泳检测并记录结果,根据Ⅰ类整合子可变区扩增产物图谱进行RFLP分型,最后进行产物的序列分析。结果PCR扩增51株鲍曼不动杆菌,Ⅰ类整合酶基因intⅠ扩增阳性45株,其中包括可变区阳性32株。并且Ⅰ类整合子的可变区扩增出的目的片段大小各不相同,有28株扩增片段约为2.3kb,2株扩增片段约为2.4kb,2株扩增片段为3kb;ISCR1扩增阳性22株,其下游扩增阳性5株,扩增片段大小约为3kb。同时携带Ⅰ类整合子和ISCR1扩增阳性菌目的片段大小约为2kb。酶切图谱显示,大小为2.3kb的Ⅰ类整合子可变区产物为同一带型;序列分析显示,大小不同的Ⅰ类整合子的可变区各片段所含的基因盒各不相同;耐药基因序列分析显示,5株ISCR1的下游扩增片段为qnrA1-ampR。同时显示Ⅰ类整合子和ISCR1两种元件以串联的形式存在于鲍曼不动杆菌中。结论Ⅰ类整合子与ISCR1大多以串联形式同时存在于骨髓炎患者感染的鲍曼不动杆菌中,该结构对耐药基因在不同鲍曼不动杆菌菌株间的水平传播可能起介导作用。     

整合子介导产生超广谱β-内酰胺酶志贺菌的多重耐药研究

目的探讨产生超广谱β-内酰胺酶（ESBL）志贺菌的耐药特性及其耐药机制。方法采用K-B法药敏试验筛选可疑产ESBL志贺菌株;通过改良三维试验、E-test试验及相对水解率测定对可疑产ESBL志贺菌进行鉴定;采用TEM、SHV、CTX-M-1组、CTX-M-2组和CTX-M-9组β-内酰胺酶通用引物进行PCR检测,并对TEM和CTX-M-9组全编码基因引物等PCR扩增产物进行DNA序列分析;对产ESBL志贺菌进行结合传递试验,供体菌和接合子用稀释法进行MIC测定。对ESBL菌株进行Ⅰ类、Ⅱ类和Ⅲ类整合子的多重PCR检测,Ⅰ类整合子可变区PCR扩增产物进行DNA测序,确定耐药基因盒的种类和数量。结果275株志贺菌中有12株为产ESBL志贺菌,其基因型为CTX-M-14和CTX-M-1组;产ESBL志贺菌结合传递试验全部阳性,其结合子只对β-内酰胺类抗生素耐药。12株ESBL志贺菌只含Ⅰ类整合子,整合子可变区含有dfrA17-aadA5耐药基因盒。结论济南地区志贺菌对β-内酰胺类抗生素的交叉耐药由质粒介导,对磺胺类和氨基糖苷类多重耐药由整合子介导。     

Variable gene cassette patterns of class 1 integron-associated drug-resistant Escherichia coli in Taiwan

This study characterized class 1 integrons in Escherichia coli in Taiwan. The stability and changes in gene cassettes inserted into integrons were also evaluated. The study included 436 clinical strains of E. coli isolated in 2002. Class 1 integrons were characterized by polymerase chain reaction and direct sequencing. Genetic localization of class 1 integrons was determined by conjugal transfer and Southern hybridization. The results indicated that 64% of E. coli isolates carried class 1 integrons. Molecular analysis revealed that the class 1 integrons harbored 13 different antimicrobial resistance gene cassettes and two unknown gene cassettes; the predominant cassettes were aadA and dfrA. Novel gene cassettes first recovered from E. coli were aacA4 and linF. Cassette arrays orfD-aacA4-catB8 and aadA1-linF were also observed. Gene cassette dfrA12-orfF-aadA2 was stable. The class 1 integron and dfrA17-aadA5 gene cassette were located on the same transferable plasmids and were capable of transmission. Therefore, the increased drug resistance of clinical isolates may be explained by antibiotic selective pressure and widespread presence of integrons. Under antibiotic selective pressure, gene cassette-mediated resistance may not be easily lost. The potential role of integrons in the uptake and dissemination of resistance genes by plasmid between species of bacteria may decrease the therapeutic effectiveness of antibiotics.     

Class 1 integrons and resistance gene cassettes among multidrug resistant Salmonella serovars isolated from slaughter animals and foods of animal origin in Ethiopia

The present study was undertaken to identify and characterize integrons and integrated resistance gene cassettes among multidrug resistant (MDR) Salmonella isolates from slaughter animals and food products of animal origin in Ethiopia. A total of 98 epidemiologically unrelated Salmonella isolates comprising 13 serovars were characterized using serotyping, phage typing, antimicrobial resistance testing and the pulsed-field gel electrophoresis (PFGE) method. Integron-PCR was used to detect the presence of class 1 and class 2 integrons in the MDR strains. The associated individual resistance gene cassettes were identified using specific PCRs and DNA sequencing. The location of the integrons was determined by Southern blot hybridization analysis. Among the Salmonella serovars, a high level of antimicrobial resistance was found to streptomycin (82.6%), tetracycline (75.5%), sulfamethoxazole (60.2%), spectinomycin (53.1%), ampicillin (42.8%), nalidixic acid (34.7%), nitrofurantoin (30.6%), trimethoprim (27.5%), gentamicin (20.4%) and ciprofloxacin (19.4%). Class 1 integrons were detected in 53.1% of the MDR isolates comprising serovars Anatum, Braenderup, Kentucky, Saintpaul and Typhimurium. Of the class 1 integron positive isolates 61.5% harboured the integron-associated gene cassettes: aadA2, aadA2+bla(PSE-1), dfrA1-aadA1 and dfrA12-orf-aadA2 (amplicon sizes 1000 bp, 1000+1200 bp, 1600 bp and 1900 bp, respectively). The chromosomally located aadA2 and aadA2+bla(PSE-1) resistance gene cassettes occurred exclusively in S. Typhimurium DT104 isolates, the other cassettes were found on large plasmids in several serovars. An aacCA5-aadA7 gene cassette array (amplicon size 1600 bp) was exclusively found in all MDR S. Kentucky strains of R type Str/SpeSmxGenNalAmpTetCipCef and this integron was shown to be chromosomally located. Results of the present study indicate that class 1 integrons carrying gene cassettes, which confer resistance to different classes of antimicrobials such as aminoglycosides, beta-lactams and trimethoprim are widespread among the MDR Salmonella serovars isolated from slaughter animals and food products of animal origin in Ethiopia indicating the important role of these genetic elements in the dissemination of multidrug resistance.     

Characterization of class 1 integrons and antimicrobial resistance in CTX-M-3-producing Serratia marcescens isolates from southern Taiwan

The objective of this study was to characterize the gene cassettes of class 1 integrons and antimicrobial resistance among CTX-M-3-producing Serratia marcescens isolates from different specimens in southern Taiwan. One hundred and twenty-two isolates (70.5%) of 173 CTX-M-3-producing S. marcescens isolates were positive for class 1 integrons, including 53.3% of blood isolates, 94.1% of urine isolates, and 87.2% of sputum isolates. No class 2 or class 3 integrons were detected in this study. By PCR with primers 5'-CS and 3'-CS for the amplification of gene cassettes regions, amplicons ranging from 0.7 to 3.0 kb in length were found in 108 (88.5%) of the 122 class 1 integron-containing isolates of CTX-M-3-producing S. marcescens isolates. Ten different types by pattern of amplicons for class 1 integrons were obtained. The Type I amplicon (46.3%) harbors two different class 1 integrons containing the gene cassette of aadA2 and aadB-catB3, respectively, and was most prevalent in the gene cassette region-positive S. marcescens isolates, followed by the Type II amplicon, which harbors one class 1 integron containing the gene cassette dfrA12-orfF-aadA2 (28.7%). Most of the S. marcescens isolates (66.7%, 8/12) harboring three different class 1 integrons (Type IV amplicon) were found in blood isolates. Class 1 integrons were conjugally transferred to recipients in 92.0% of S. marcescens harboring two different class 1 integrons containing the gene cassettes aadA2 and aadB-catB3, respectively. The transfer rate of class 1 integron carrying dfrA12-orfF-aadA2 was detected in 77.4% of S. marcescens isolates. The results showed that all those isolates with conjugative transfer of integrons carried their class 1 integrons on the conjugative plasmids.     

天津地区福氏志贺菌整合子携带及耐药性研究

目的了解天津地区不同年份福氏志贺菌血清型分布,整合子携带情况,耐药性及其变化趋势。方法采用血清学方法对天津地区不同年份分离的56株福氏志贺菌进行分型;PCR扩增整合子整合酶及可变区,并测序;采用K-B法测定其对16种抗菌药物的敏感性。结果 1981~1983年分离株福氏志贺菌1类整合酶阳性率为87.50%(28/32),其中27株可变区含氨基糖苷类药物耐药基因aadA;2009~2011年分离株福氏志贺菌1类整合酶阳性率为91.67%(22/24),可变区及3’末端扩增均阴性。1981~1983年分离株福氏志贺菌2类整合酶均阴性;2009~2011年分离株福氏志贺菌2类整合酶阳性率79.17%(19/24),可变区含dfrA1、sat1、aadA1,介导对甲氧苄啶、链丝菌素和链霉素耐药。有17株福氏志贺菌1、2类整合酶均阳性。1981~1983年分离株福氏志贺菌对四环素、链霉素、氯霉素及甲氧苄啶/磺胺甲噁唑耐药率高,多重耐药率为65.63%;2009~2011年分离株福氏志贺菌对氨苄西林、链霉素、甲氧苄啶/磺胺甲噁唑、哌拉西林和四环素耐药率高,多重耐药率为83.33%。结论 1、2类整合子广泛存在于天津地区福氏志贺菌中,其中1类整合子3’保守末端可能缺失或存在变异。2009~2011年分离株福氏志贺菌对抗菌药物耐药性较1981~1983年分离株增强,多重耐药率增高。福氏志贺菌优势血清型发生转变,血清型分布呈现多样化。     

多重抗药大肠杆菌中Ⅰ型整合子的分布与抗药关系

目的检测多重抗药大肠杆菌中检测Ⅰ型整合子的携带率及其对细菌抗药性的影响。方法用大肠杆菌显色．培养基分离奶牛场、猪场和肉鸡场的565株大肠杆菌,测定其对15种抗生素的MIC;用PCR扩增其中91株多重抗药性（最少耐3种抗生素）菌株的Ⅰ型整合酶基因及抗药基因盒。结果565株大肠杆菌仅对头孢噻呋、阿米卡星和硫酸粘菌素的抗药率低于50％,其余药物均表现出较高抗药率,且几乎所有细菌都对3种以上抗生素具有抗性。91株多重抗药大肠杆菌Ⅰ型整合子的平均检出率为83．52％,大多数整合aadA基因和dhfr基因,抗药性基因携带率为43．42％。结论Ⅰ型整合子对细菌多重抗药性的产生和传播起着重要作用,是临床细菌多重抗药性监测在基因水平的重要指标。     

Phenotypic and genotypic characterization of Salmonella enterica recovered from poultry meat in Tunisia and identification of new genetic traits

Thirty-seven Salmonella enterica isolates obtained from poultry meat in Tunisia were included in this study for characterization of antibiotic resistance mechanisms. High percentages of resistance were detected to ampicillin, sulfonamides, tetracycline, nalidixic acid, and streptomycin (32.4%-89.2%), and lower percentages to amoxicillin-clavulanic acid, kanamycin, amikacin, trimethoprim-sulfamethoxazol, and chloramphenicol (2.7%-18.9%). All strains showed susceptibility to ceftazidime, cefotaxime, gentamicin, and ciprofloxacin. Class 1 integrons were detected in 30% of Salmonella isolates, and four different gene cassette arrangements were detected, including genes implicated in resistance to aminoglycosides (aadA1 and aadA2) and trimethoprim (dfrA1). Four different Pc variants (PcW, PcH1, PcH1(TTN-10), PcW(TGN-10)) with inactive P2 have been found among these isolates. Integron-positive isolates were ascribed to eight different serotypes. A Salmonella Schwarzengrund isolate harbored a new class 1 integron containing the qacH-dfrA1b-aadA1b-catB2 gene cassette arrangement, with the very unusual PcH1(TTN-10) promoter, which has been registered in GenBank (accession no. HQ874651). Different plasmid replicon types were demonstrated among integron-positive isolates: IncI1 (8 isolates), IncN (8), IncP (2), IncFIB (2), and IncFII (2). Ten different pulsed-field gel electrophoresis profiles were detected among the 11 integron-positive isolates and 8 different sequence types were identified by multilocus sequence typing, one of them (registered as ST867) was new, detected in 3 Salmonella Zanzibar isolates. A high diversity of clones is observed among poultry Salmonella isolates and a high proportion of them show a multiresistant phenotype with very diverse mobile genetic structures that could be implicated in bacterial dissemination in different environments.     

Molecular diversity of class 1 integrons in human isolates of Aeromonas spp. from southern Taiwan

This work studies antimicrobial resistance and class 1 integrons of Aeromonas spp. in human isolates from southern Taiwan. PCR amplification and DNA sequence analyses were performed to characterize the gene cassette regions of the class 1 integron in 204 isolates of Aeromonas hydrophila, 36 isolates of A. sobria, 23 isolates of A. veronii, and 4 isolates of A. caviae. By using Southern hybridization with an intI1 probe to determine the presence of class 1 integrons in the 9 isolates of Aeromonas spp. harboring plasmid DNA, only 2 isolates, one A. veronii AV69 harboring 176-kb plasmid DNA, and one A. hydrophila AH207 harboring 149-kb plasmid DNA were identified. A conjugation experiment was carried out with 2 isolates of A. veronii AV69 and A. hydrophila AH207. Only one transconjugant of Escherichia coli AH207, containing 149-kb plasmids obtained from A. hydrophila AH207, was identified. ERIC-PCR analysis was performed to analyze the genetic relatedness in all isolates of Aeromonas spp. that carry class 1 integrons. The results of cluster analysis in this experiment revealed that none of these isolates were clonal, which may indicate that they were not related to the outbreak. Among the 267 isolates tested, class 1 integrons were detected in 37 isolates (13.9%) of Aeromonas spp. from humans. No class 2 or class 3 integrons were detected in this study. Gene cassette structures were identified in 30 (81.0%) of 37 isolates of Aeromonas spp. containing class 1 integrons. The gene cassette of dfrA12-orfF-aadA2 was the most prevalent in the gene cassette array (16.0%), followed by arr3-aacA4 (13.3%) and dfr2d-catB3-aadA1 (10.0%). Four novel arrays of gene cassettes were also identified, namely, dfr2d-catB3-aadA1, aadA1-aac(6')-II, aadA4a, and aac(6')-II-blaOXA-21-catB3. This is the first report of Aeromonas spp. isolates from humans.