Use of plasmid profile typing for surveillance of Salmonella enteritidis phage type 4 from humans, poultry and eggs.

Plasmids were found in 1022 of 1089 (94%) of drug-sensitive strains of Salmonella enteritidis phage type 4 from humans (sporadic and outbreak cases), poultry (chickens) and eggs in England and Wales in the 5-year period 1988-92 and 25 plasmid profile patterns were identified. Strains characterized by a single plasmid of 38 MDa predominated (= plasmid profile type SE 38), comprising over 90% of isolates from humans, 70% from poultry and 92% from eggs. Eleven profile types were identified in strains from humans, 21 in strains from poultry and 3 in strains from eggs. Eight of the 11 patterns identified in human isolates were found in strains from poultry and 2 in strains from eggs. In contrast 15 patterns seen in poultry were not found in strains from humans. Four percent of strains from humans and 13% from poultry did not carry the 38 MDa plasmid but all strains from eggs were found to carry this plasmid. The second most common profile type in strains isolated between 1981 and 1988 was not identified in strains isolated from 1988-92. It is concluded that plasmid profile typing is a useful method for rapid differentiation within phage type 4 of S. enteritidis but that methods which can discriminate within the predominant profile type, SE 38, are now required.     

Gentamicin-resistant Salmonella typhimurium phage type 204c: molecular studies and strain diversity in a putative bovine outbreak

As part of the investigation of a putative bovine outbreak, 13 isolates of Salmonella typhimurium phage type 204c were subjected to plasmid analysis. Plasmid profiles suggested that several distinct strains were involved and these observations were supported by minor variations in antibiotic resistance pattern. Restriction enzyme fingerprinting and conjugational segregation of the plasmids confirmed these findings. Although 12 of the 13 isolates were resistant to gentamicin, resistance was conferred by 4 distinct plasmids; 3 of these belonged to Inc I and were distantly related on the basis of restriction fingerprints and the fourth was a resistant derivative of the 60 MDa S. typhimurium serotype-specific plasmid. The molecular evidence refuted the hypothesis that geographical and temporal clustering of these gentamicin-resistant isolates could be explained on the basis of a single epidmiological episode.     

Plasmid profiles of antibiotic-resistant Shigella dysenteriae types 2, 3, 4, 6 and 7 isolated in Ethiopia during 1976-85.

Plasmid profile analysis by agarose gel electrophoresis was carried out on 37 drug-resistant strains of Shigella dysenteriae types 2, 3, 4, 6 and 7. These strains were collected between 1976 and 1985 in Addis Ababa, Ethiopia. The plasmid profile of S. dysenteriae type 2 strains with R-type CSSuT did not show middle-sized plasmids likely to code for CSSuT resistance. All strains contained a large plasmid of about 120 megadaltons (MDa), and a cryptic plasmid of about 2.2 MDa. The plasmid profiles of S. dysenteriae type 3 with R-types ACSSuT, SSuT and SSu showed a 4.2 MDa SSu-determinant, which was demonstrated in Escherichia coli K12 recipients resulting from triparental crosses. The ACT determinant in S. dysenteriae type 3 with R-type ACSSuT is probably chromosomally mediated. Cryptic plasmids of about 3.0 and 2.2 MDa were found in all S. dysenteriae type 3 isolates. The 4.2 MDa plasmid featured prominently in the plasmid profiles of S. dysenteriae types 4, 6 and 7 with R-types SSuT and SSu. However, this plasmid was not mobilizable by triparental crosses. There was a relative paucity of transferable plasmids in non-Shiga bacillus isolates. However, incompatibility group N plasmids, coding for tetracycline resistance, were detected.     

Salmonellosis in pigmy hogs (Sus salvanius)--a critically endangered species of mammal

The pigmy hog (Sus salvanius) is the smallest and the rarest wild suid in the world. This species is on the verge of extinction and the World Conservation Union has rated it among the most endangered of all mammals. This paper reports the investigation into an outbreak of salmonellosis among captive pigmy hogs at the Research and Breeding Centre of the pigmy hog conservation programme, Guwahati, Assam, India. Of 75 pigmy hogs (38 males and 37 females) maintained at the Centre, seven (9.3%) died within five days. The causative organism associated with the outbreak was identified as Salmonella Typhimurium (syn. S. enterica serovar Typhimurium). All the isolates of S. Typhimurium belonged to phage type DT193. The isolates harboured multiple plasmids. Five isolates harboured four (65.0 MDa, 4.2 MDa, 3.0 MDa, 1.3 MDa), while two isolates carried three plasmids (65.0 MDa, 4.2 MDa, 3.0 MDa). All strains showed resistance to amikacin, ampicillin, streptomycin and sulfamerazin; five strains were resistant to oxytetracycline and trimethoprim. All the strains were sensitive to chloramphenicol, ciprofloxacin, enrofloxacin and gentamicin. All seven isolates of S. Typhimurium were found to harbour stn, sopB and pefA genes. However, none of them was found to carry sefC and sopE genes.     

Plasmid content, auxotype and protein-I serovar of gonococci isolated in the Gambia

Twenty-nine strains of penicillinase-producing Neisseria gonorrhoeae (PPNG) and 30 non-penicillinase-producing strains, all isolated in the Gambia, were characterized in terms of their plasmid content, auxotype and protein-I serovar. Sixty-two per cent of the PPNG strains contained the 3.2 MDa penicillinase-coding plasmid, and 38% had the 4.4 MDa plasmid. All the PPNG strains contained the 2.6 MDa cryptic plasmid but lacked the 24.4 MDa conjugative plasmid. In contrast, 46.7% of the non-PPNG strains harboured only the cryptic plasmid while 16.7% contained both the cryptic and conjugative plasmids. Seventeen per cent of the non-PPNG strains contained the conjugative plasmid only and 20% lacked plasmids. The PPNG and non-PPNG strains also differed in terms of their protein-I serovar. Eighty-six per cent of the PPNG strains belonged to serogroup 1 A, whereas the majority (60%) of non-PPNG strains belonged to serogroup 1 B. There was no significant difference in the auxotypes of the PPNG and non-PPNG strains, with both groups consisting predominantly of prototrophic and proline-requiring strains, with a minority of strains requiring arginine. When the 59 strains were each characterized in terms of their combined plasmid profile, auxotype and serovar, 39 different combinations were noted, which indicates the heterogeneous nature of the gonococcal population found in the Gambia.     

Simultaneous infection with three different S. enteritidis strains in a nursing home resident

A culture taken from a nursing home resident as part of a S. enteritidis outbreak was found to have a mixed infection due to three different strains of S. enteritidis. One of the three strains belonged to phage type (PT) 4, one to PT6 and one reacted with phages but did not conform to any typing scheme (RDNC). All three strains had the 38.9 megadaltons (MDa) plasmid found in the isolates from the outbreak-related cases, in addition the PT6 and RDNC strains harboured a 69.9 MDa plasmid. The importance of phage typing and plasmid analysis for S. enteritidis strain characterization and their epidemiologic and bacterial significance are discussed.     

深圳市龙岗区沙门菌质粒图谱和耐药性分析

目的了解深圳市龙岗区沙门菌的质粒图谱分布和耐药性。方法利用快速小量质粒提取技术及电泳图谱方法,对深圳市龙岗区2003—2007年食物中毒及伤寒、副伤寒疫情中分离的67株沙门菌进行质粒图谱分析并做药敏试验,药敏试验采用改良K—B法。结果质粒阳性率为67．2％（45／67）。质粒携带株每株1—8个不等数量的质粒,相对分子质量大小为1．1。54．2kb,质粒条带数以1～3条居多,未发现耐药谱与质粒特征有关。沙门菌对氟哌酸、氯霉素、氨基糖苷类及第3代头孢类抗生素完全敏感,对青霉素、苯唑西林完全耐药,对头孢氨苄的耐药率高达41．8％,对氨苄西林、羧苄西林、哌拉西林、四环素和复方新诺明的耐药率分别为37．3％、37．3％、25．4％、25．4％和3．0％;沙门菌多重耐药率为13．0％。结论深圳市龙岗区人群携带的沙门菌耐药情况比较普遍;检测沙门菌质粒阳性率不高,质粒特征与耐药谱无关。     

Drug resistance and plasmid profile of shigellae in Taiwan.

One hundred and twenty-eight shigella strains isolated from newborn and infant human faecal specimens at Kaohsiung Medical College Hospital in Taiwan were serogrouped, serotyped and examined for drug-resistance patterns and for the presence of plasmids. Forty-seven per cent of the isolates were found to belong to the Shigella sonnei serogroup, 41% to the S. flexneri group, 9% to the S. boydii group and 3% to the S. dysenteriae group. The serotype with the greatest number of strains was S. sonnei I. (29%) followed by S. flexneri 1 (27%). Each strain was tested for resistance to 11 antimicrobial agents. Eight-eight per cent of the strains were resistant to tetracycline, 87% to chloramphenicol, 84% to streptomycin, 52% to ampicillin, 25% to nalidixic acid, 29% to kanamycin, 11% to cephalothin, 11% to neomycin, 10% to cotrimoxazole, 1% to amikacin and none to gentamicin. The most prevalent resistance pattern was ApCmSmTc (28%). Clinical isolates demonstrating multiple resistance were found to harbour a large transmissible plasmid of 45-75 MDa while isolates without multiple resistance did not. Two large virulence plasmids of 123 and 110 MDa were found in 12 strains of S. flexneri and 4 strains of S. sonnei phase I. Small plasmids of 4.5, 4.2, 3.5, 2.8, 2.5. 2.0 and 1.5 MDa were also present in all strains. These small plasmids were species specific and can be used as marker plasmids to identify species.     

Identification of multiple-resistance (R) and colicinogeny (Col) plasmids in an epidemic Salmonella agona serotype in Rio de Janeiro.

A Salmonella agona strain has caused a hospital outbreak of gastroenteritis in a pediatric unit in Rio de Janeiro. It bears two plasmids, a small (6.5 MDa molecular weight) plasmid coding for type B colicin production and a larger one (36 MDa molecular weight) determining resistance to ampicillin, gentamicin, kanamycin and trimethoprim-sulphamethoxazole. The R-plasmid, but not the Col-plasmid, is self-transferable to a Escherichia coli recipient strain. Curing for the R-plasmid was achieved by treatment with 0.05% SDS followed by incubation at 44 degrees C. It has not been possible to cure the S. agona strain for its Col-plasmid.     

Egg-related Salmonella enteritidis, Italy, 1991

In recent years, Salmonella enteritidis has become an increasingly important public health problem in Italy. In some parts of the country, the fraction of total human salmonella isolates accounted for by S. enteritidis has risen from 3-4% in the mid-1980s to more than 30% in 1990. Between 1990 and 1991, the number of reported S. enteritidis outbreaks increased more than sixfold. The 33 outbreaks reported in 1991 occurred in seven contiguous regions in northern and central Italy and were clustered in time between June and October; in the majority, products containing raw or undercooked shell eggs were implicated. Five of the egg-related outbreaks that occurred within a 30 kilometre radius over a 7-week period were investigated in detail. A phage type 1 strain containing a 38·9 MDa plasmid appeared responsible for three of the outbreaks, while in the remaining two a phage type 4 strain, also with a 38·9 MDa plasmid was isolated. Efforts are being made to enhance epidemiological surveillance and laboratory evaluation, and the use of pasteurized eggs has been recommended for high-risk populations.     

The emerging strains of Shigella dysenteriae type 2 in Bangladesh are clonal

A total of 113 strains of Shigella dysenteriae type 2 isolated from patients attending the Dhaka diarrhoea treatment centre of ICDDR,B: Centre for Health and Population Research during the period 1999-2004 were studied. Serotype of the isolates was confirmed using commercially available antisera. Except for arabinose fermentation, all the strains had similar biochemical reactions. More than 60% of the strains were sensitive to commonly used antibiotics; only 6% (n=7) of the strains were resistant to nalidixic acid, and none of the strains were resistant to mecillinam and ciprofloxacin. All strains were invasive as demonstrated by the presence of a 140 MDa plasmid, ial, sen and ipaH genes, Congo Red absorption ability and by the Sereny test performed on representative strains. Plasmid patterns were heterogeneous but more than 50% of strains were confined to a single pattern. All strains possessed a 1.6 MDa plasmid and 87% of the strains contained a 4 MDa plasmid. Middle-range plasmids (90 MDa to 30 MDa) present in 36% of the strains were not associated with antibiotic resistance. All the strains were clustered within a single type with four subtypes by pulsed-field gel electrophoresis while ribotyping patterns of all the strains were identical.     

Insertion sequence IS200 fingerprinting of Salmonella typhi: an assessment of epidemiological applicability.

When Pst I-generated digests of genomic DNA from each of the type strains of 49 of the Vi phage types of Salmonella typhi were probed with a PCR-amplified IS200 gene probe, all strains were found to possess at least 11 IS200 elements carried on fragments in the range 24.2-1.2 kb. Fourteen fingerprints were identified but two patterns designated IS200Sty1 and IS200Sty2 predominated. In one strain, a plasmid-mediated IS200 element was identified. When IS200 fingerprinting was applied to epidemiologically-unrelated strains of S. typhi isolated in Ecuador, 3 patterns were identified in 10 strains belonging to 9 different phage types. It is concluded that Vi phage typing remains the method of choice for the primary differentiation of S. typhi but that IS200 fingerprinting may be of limited use in laboratories which do not have access to phage typing.     

Molecular relationship among Salmonella dublin isolates identified at the Center for Enterobacteriaceae of Palermo during the years 1971-85

A molecular epidemiological study was carried out on 60 Salmonella dublin isolates identified at the Southern Italy Enterobacteriaceae Center between 1971 and 1985. These included 23 isolates from children with diarrhoea in Palermo obtained during 1984. All isolates from the outbreak of gastroenteritis in children were resistant to chloramphenicol and streptomycin and harboured two plasmids of 50 MDa and 3 MDa molecular weight, whereas the majority of the isolates identified before 1984 were susceptible to these antibiotics and carried only a 50 MDa molecular weight plasmid. Four S. dublin strains successively identified from cattle (Palermo, Foggia, Portici) and from a child (Palermo) were shown to possess similar antibiotic resistance patterns and plasmid profiles to S. dublin isolates from the outbreak of gastroenteritis in children. The 50 MDa plasmid was shown to be associated with virulence in mice, while it was not possible to assign any genetic function to the 3 MDa plasmid.     

Molecular characterization of trimethoprim resistance in Shigella sonnei in Sicily

During the 3-year period 1985-7, all strains of Shigella sonnei isolated in Catania, Sicily, showed a high level of resistance to trimethoprim (Tp) which was invariably associated with resistance to other antibiotics. Plasmid analysis showed 18 different electropherotypes: 35 of 37 strains harboured a plasmid of 70 Megadaltons (MDa), and 29 of 37 strains a plasmid of 130 MDa. Restriction endonuclease fingerprinting of purified 70 MDa plasmid DNA from different strains demonstrated that these plasmids were similar but not identical. In some strains with transferable Tp resistance, DNA hybridization analysis demonstrated the presence of gene coding for the production of dihydrofolate reductase (DHFR) type V. In contrast, there was no detectable hybridization with DNA probes specific for genes coding for DHFR types I, II and IV. This is the first report of the DHFR type V gene outside Sri Lanka.     

Phage type conversion in Salmonella enterica serotype Enteritidis caused by the introduction of a resistance plasmid of incompatibility group X (IncX)

The plasmid pOG670, a 54 kb, conjugative plasmid that specifies resistance to ampicillin and kanamycin and belonging to the incompatibility group X (IncX), was transferred into 10 isolates of Salmonella enterica serotype Enteritidis belonging to 10 different phage types (PT1, 2, 3, 4, 8, 9, 9b, 10, 11 and 13). Acquisition of the plasmid by these strains did not result in the loss of any resident plasmids but resulted in phage type conversion in 8 of the 10 strains (PT1, 2, 4, 8, 9, 9b, 10 and 11). The observed changes in phage type were found to result from the loss of sensitivity to 3 of the 10 typing phages used (phages 3, 5 and 7). Where the conversion resulted in a change to a defined phage type, both the new and original PTs belonged to the same, previously described, evolutionary lines. Enteritidis PTs 1, 4 and 8, commonly associated with poultry world-wide, were converted to PTs 21, 6 and 13a respectively. The results indicate a different route for phage type conversion Enteritidis from others reported in the literature and, although IncX plasmids are not normally present in PT8 or PT13a, may suggest a possible mechanism/link connecting these phage types.     

Subdivision of Salmonella enteritidis phage types by plasmid profile typing

Differentiation of Salmonella enteritidis by plasmid profile typing has been compared to differentiation by phage typing. Examination of the type strains of the 27 S. enteritidis phage types showed that only 11 profile patterns could be identified. Moreover, two profile patterns were found in 15 of the type strains, including those of the two most common phage types in Britain, types 4 and 8. On this basis, plasmid profile typing is not as sensitive as phage typing for the primary subdivision of S. enteritidis. When differentiation of 534 strains of the 27 phage types was attempted using plasmid profiles, variation in pattern suitable for epidemiological subdivision was found in 13 phage types and there were 9 profile patterns in strains of phage type 4. Plasmid profile typing can, therefore, be regarded as an effective adjunct to phage typing for the subdivision of S. enteritidis.     

Diversity of plasmids in Staphylococcus saprophyticus isolated from urinary tract infections in women

A group of 150 Staphylococcus saprophyticus strains isolated from urinary tract infections in women were included in this study. Antimicrobial susceptibility tests showed that these isolates were sensitive to most antimicrobial agents. All strains were sensitive to penicillin, cephalothin, gentamicin, kanamycin, trimethoprim and nitrofurantoin. Resistance to tetracycline was present in 10.6% of the strains, to chloramphenicol in 4%, to erythromycin in 1.3% and to streptomycin in 1.3%. All strains were resistant to cadmium chloride as well as to novobiocin and nalidixic acid. Plasmid analysis showed that 82% of the strains harboured plasmids, some of them with complex plasmid profiles. Most plasmids were considered to be cryptic, although antibiotic resistance plasmids were identified in 18 isolates. Tetracycline resistance was encoded by a plasmid of c. 2.8 MDa, chloramphenicol resistance by a plasmid of c. 2.9 MDa and erythromycin resistance by a plasmid of c. 1.6 MDa. Streptomycin resistance could not be linked to the presence of any specific plasmid. Plasmid profiling seemed to be a good method for differentiating among S. saprophyticus strains.     

Molecular epidemiology ofSalmonella enteritidis

Sixteen strains ofSalmonella enteritidis isolated in 1991 from 13 unrelated poultry-associated sources, 7 strains from 2 community outbreaks, and 18 human sporadic isolates were investigated by phage typing, analysis of rRNA gene restriction patterns (ribotyping) and plasmid profiles. Four different phage types and 10SphI patterns were found, whereas plasmids were identical in all but 4 isolates. Only one ribotype (RT A) occurred among both human and avian strains. This particular ribotype was also responsible for the two outbreaks investigated, suggesting that such strains may be of special significance for the increase ofS. enteritidis infections.     

Characterization of resistance plasmids and carried phages in an epidemic clone of multi-resistant Salmonella typhimurium in India.

Hospital outbreaks of severe gastroenteritis caused by multi-resistant Salmonella typhimurium have occurred in a number of cities throughout India since 1977. The strains involved belong to phage types 66 or 122, or are untypable; the latter are derived from types 66 or 122 by acquisition of one or more of a number of temperate bacteriophages. Types 66 and 122 are closely related and react with the same phages of the S. typhimurium typing scheme. A plasmid belonging to compatibility group F1me encoding resistance to ampicillin, chloramphenicol, kanamycin, streptomycin, sulphonamides, spectinomycin, tetracyclines, gentamicin and trimethoprim (R-type ACKSSuSpTGTm) is present in all of the multi-resistant strains. Several other plasmids have been identified including an SSu resistance determinant, a group I2 transfer factor and an R factor coding for resistance to kanamycin, streptomycin and sulphonamides which is compatible with plasmids of all the standard compatibility groups. These plasmids are only present in a proportion of the strains examined. Examination of strains from other sources has identified a paediatric hospital outbreak in Saudi Arabia and a number of sporadic infections in Great Britain which have been caused by the same organisms. These studies show that, despite differences in phage type and plasmid content, this group of strains belongs to a single clone which has become widespread in India with some extension to other countries.     

Electropherotyping of plasmid DNA of different serotypes of Shigella flexneri isolated in Bangladesh

One hundred and twenty-five Shigella flexneri strains, isolated during January-December 1984, at the Dhaka treatment centre of the International Centre for Diarrhoeal Disease Research, Bangladesh, were serotyped using absorbed rabbit antisera specific for all type- and group-factor antigens, as well as a group of ten mouse and rat monoclonal antibodies. Electropherotypes of the plasmid deoxyribonucleic acid (DNA) were also determined. S. flexneri 2a was the predominant serotype followed by 3b, 1a, and 2b. The recently described E1037 antigen was also found in three strains of S. flexneri serotype 6. Electropherotyping of the plasmid DNA showed that three plasmids of approximately 140, 2.7, and 2 megadalton (MDa) were present, respectively, in 97, 97 and 94% of the 125 strains. Additional plasmids of various other sizes were also present in different serotypes except in serotype 2a. The additional plasmids again appeared to be specific for that particular serotype. For example, all 12 strains of S. flexneri 2b harboured an additional plasmid of approximately 1 MDa. Thus, electropherotyping of plasmid DNA of different serotypes of S. flexneri might be useful to differentiate S. flexneri from other species of Shigella and in identifying different serotypes of S. flexneri. Therefore, the common plasmids, plus the additional plasmids, could be used to identify epidemic, as well as sporadic, subclones of S. flexneri strains.