Identification of two plasmids determining resistance to tetracycline and to erythromycin in group D streptococcus

Summary Two circular DNA plasmids designated Tet and Ero, distinguishable by their molecular weights (64.5 and 17.6×10⁶ respectively) and their guanine plus cytosine (G+C) content (36.5 and 34.5% respectively) were identified in Streptococcus faecalis strain KR, resistant to tetracycline and erythromycin. Loss of resistance to tetracycline, to erythromycin and to both antibiotics simultaneously in independently isolated cured derivatives corresponds to the loss of the Tet, Ero, and of both plasmids respectively. In the singly resistant strains we estimated that 3 and 6 copies of the Tet and Ero plasmids respectively are present per chromosomal genome equivalent.Abbreviations form I supercoiled circular DNA - form II relaxed circular DNA - form III linear double-strand DNA - Tet tetracycline - Ero erythromycin - R resistant - S sensitive     

Repetition of tetracycline resistance determinant genes on R plasmid pRSD1 in Escherichia coli.

Summary The 30 megadalton (Mdal)-conjgaative, fi^- plasmid pRSD1 determines inducible tetracycline resistance (Tc) in Escherichia coli. As shown by restriction analysis, a 3.5 Mdal-EcoRI fragment of pRSD1 spliced into the small plasmid pRSD2124 comprises the entire Tc determinant (tet) region. A restriction map of pRSD1 is presented which includes the location of the tet region and of an underwound loop not related to Tc (Burkardt et al., 1978). Selective amplification of tet genes is demonstrated by three lines of evidence. (i) The resistance level of cell harbouring pRSD1 increases approximately tenfold by induction with 10g/ml of tetracycline. Further groth in the presence of 100 g/ml of the drug (tet-racycline stress) selects for cells with even higher resistance levels (about 300 g/ml) in rec ⁺ cells. In a recA strain, a smaller proportion of cells attains these high resistance levels suggesting the involvement of host recombination. (ii) Electron micrographs of pRSD1-DNA isolated from tetracycline-stressed cells reveal a heterogeneous population of circular DNA molecules ranging between 1.7 and 21.6 m. The distribution of contour lengths shows a discrete pattern ascribed to the presence of autonomous single-and multiple-copy Tc determinants and to intact plasmids containing zero to six tet regions in tandem repeats. (iii) This interpretation is supported by heteroduplex and restriction analyses which demonstrate the presence of multiple copies of the 3.5 Mdal-element encompassing the tet region in pRSD1 molecules selected by tetracycline stress. It has been concluded that gene amplification leading to tandem repetition of the tet region ensues in pRSD1. Such plasmids confer increased tetracycline resistance and can, therefore, be selected by high doses of the drug.     

Non-random distribution of transduction termini in transductants from the integrated R plasmid, R100-1

Summary Tra ⁺and tra ^–derivatives of drug resistance plasmid, R100-1, were isolated by phage P1 from an Hfr donor with integrated R100-1 and then analyzed by complementation tests with tra ^–point mutants of Flac. Tra ⁺derivatives of R100-1 carrying tetracycline resistance alone and those carrying all six drug-resistance genes could support transfer of tra ^–point mutants of Flac except Flac traJ, whereas all of tra ^–derivatives of R100-1 failed to complement any one of tra ^–point mutants of Flac. This suggests that these tra ^–derivatives of R100-1 carrying tetracycline resistance gene are deleted for all the transfer genes impaired in the Flac point mutants tested. We assume a hot point, probably a specific base sequence similar to an IS element, at the left of the tetracycline gene (Fig. 1) becomes a transduction terminus in transduction of the integrated R100-1 by phage P1. Complementation analysis of tra ^–derivatives carrying five resistance genes except the tetracycline gene led us to a supposition that a gene(s), probably analogous to traJ of the F plasmid, is located on R100-1 near the tetracycline gene which plays an important regulatory role for self-transfer as well as for the complementation of tra ^–Flac mutants.     

Specificity of bacteriophage Mu excision

Summary To study the excision of bacteriophage Mu at the DNA sequence level, the Mu-derived phage placMu3 was transposed to the transcribed but non-translated leader region of a plasmid-borne tetracycline (tet) resistance gene. Revertants (excision products) were then selected by Tet⁺ restoration of Tet⁺ and characterized. Of 21 independent Tet⁺ revertants, 17 contained simple deletions of most or all of placMu3, while the other four contained more complex rearrangements in which one end of placMu3 had been transposed, and most of the prophage had been deleted. The deletion endpoints were found in short direct repeats in each of the complex rearrangements and in 11 of the 17 simple deletion excisants. The results suggest models of slipped mispairing of template and nascent DNA strands facilitated by proteins of the Mu transposition machinery.     

Analysis of the reduction in expression of tetracycline resistance determined by transposon Tn10 in the multicopy state

The tet genes of transposon Tn10 have been mapped in a 2,200 bp DNA sequence by analysing deletion and Tn5 insertion mutations. When the tet genes were present on multi-copy plasmids the level of resistance expressed was about ten-fold lower than that determined by a single copy of Tn10 in the E. coli chromosome. The 36K tet protein known to be encoded by R100 in E. coli minicells was not detected when they harboured a multicopy tet plasmid. However, normal high levels of resistance were expressed when the tet genes were recombined into the host chromosome as part of a lambda lysogen, showing that the multicopy effect was phenotypic. Most of the Tn5 insertions and deletions in tet which caused Tc^s mutations also prevented expression of high level Tc^r from a chromosomal Tn10 element present in the same cell. Only those insertions in the promoter-proximal 90–130 bp of a 1,275 bp HindII fragment known to carry the gene encoding the 36K tet protein did not reduce the single copy Tn10 resistance level.A gene fusion system that results in the constitutive synthesis of -galactosidase from a tet promoter has been used to assay tet repressor activity. The basal (uninduced) -galactosidase level in cells carrying multicopy tet plasmids was 10–20 fold lower than those carrying a single copy. The tet:: Tn5 mutants defective in the trans-dominant multicopy effect still made normal amounts of tet repressor showing that repressor overproduction was not responsible for this effect. In addition a repressor-defective constitutive mutant did not exhibit a higher resistance level when located on a multicopy plasmid vector. We postulate that a regulatory mechanism recognises the amino-terminus of the tet structural gene product when attempts are being made to overproduce the protein and prevents further translation.     

Development of a Tn10 Vehicle for Insertion Mutagenesis in Rhizobium

A transposon Tn10 vehicle was developed using a self transmissible (Tra⁺) plasmid pRK2013 having narrow host range ori of replication (ColEl). The construct pSA10-3 carrying Tn10 was useful in efficiently transferring transposon Tn10 from E. coli into various rhizobia. The ColEl replicon conferred suicidal property to vector in Rhizobium background where it falls to replicate stably. Thus this plasmid can be employed to cause independent insertion mutations in rhizobia by Tn10 transposition. The frequency of tetracycline resistant colonies of Rhizobium (Tn10 mutants) was approximately 10⁵ folds higher than the spontaneous Tet^R mutants. Reversion frequency of these mutants was less than 10^?8 indicating adequate stability of Tn10 mutations.     

Nucleotide sequence of class D tetracycline resistance genes from Salmonella ordonez

Summary Plasmid pIP173, isolated from Salmonella ordonez strain BM2000, confers resistance to tetracycline and a number of other antibiotics. We determined the nucleotide sequence of the pIP173 tetR repressor and tetA resistance genes. The pIP173 tetR gene is essentially identical to the class D tetR gene from plasmid RA1. The pIP173 tet genes are flanked by directly repeated copies of the insertion sequence IS26. Interestingly, the 3 end of the tetR gene, encoding the C-terminal 16 amino acids of the TetR protein, extends into the flanking IS26 sequence. The relationships between the class A, B, C, and D TetA sequences parallel the relationships between the corresponding TetR sequences; class D is more closely related to class B than to either class A or C. Overall, the four TetA sequences show 38% identity and 57% similarity.     

Polarity of localised conversion in Streptococcus pneumoniae transformation

Summary Localised conversion in pneumococcal transformation is a process that spans a few nucleotides when the 5-ATTAAT/3-TAAGTA configuration occurs at the pairing step. It was first observed in two-point crosses between an amiA mutation (amiA36) carrying this sequence and other closely linked mutants of the locus. The yield of the amiA resistance allele conversion to wild type is 20%. In order to characterize this process, which differs from longpatch conversion by the length of DNA repair, gene requirements and sequence specificity, we devised expreriments to detect the reciprocal conversion, AmiA⁺ to AmiA^r. For this purpose we examined the suppressibility by a pneumococcal informational suppressor of several nonsense mutations at the locus. Amber (UAG) and ochre (UAA) mutations are suppressed whereas UGA is not suppressed. In this genetic background, where amiA36 is partly suppressed, it was possible to select for double mutants in a cross between amiA36 and a closely linked non-suppressible marker. Direct isolation of such double mutants was also performed without any screening in crosses between amiA36 and the same linked marker in cloned DNA. The frequency of double mutants was very low (1/175) suggesting that there is no conversion of wild-type to mutant alleles. Thus conversion is a polarized process changing specifically A to C.     

Induction of ?2 frameshift mutations within alternating GC sequences by carcinogens that bind to the C8 position of guanine residues Development of a specific mutation assay

Summary Using a forward mutation assay we have previously found that N-2-acetylaminofluorene (AAF), a strong chemical carcinogen, induces a majority of frameshift mutations located at specific sequences called mutation hot spots. Among these hot spot sequences, the NarI sequence (GGCGCC), is specific for –2 frameshifts (GGCGCC) GGCC). Interestingly, these frameshift mutations occur independently of a functional umuDC locus. Being interested in elucidating this mutation pathway we have developed a reversion assay that is specific for this class of mutations. The assay is based on the reversion of a +2 frameshift mutant of plasmid pBR322 from tetracycline sensitivity to tetracycline resistance. It is shown that only true reversion events lead to tetracycline resistance. The carcinogen AAF induces this reversion event at a frequency that is increased four- to fivefold over the background frequency. A series of chemical carcinogens which, like AAF, bind covalently to the C8 position of guanine, are compared for their efficiency to induce this specific mutation event. Large variations in the mutagenic efficiency of these chemicals are observed and discussed in terms of the anti/syn conformation of the carcinogen-modified guanine residue. Based on this test, we describe a convenient spot assay that this presently used in our laboratory to isolate Escherichia coli mutants affected in this mutation pathway.     

Establishment of Escherichia coli cells with an integrated high copy number plasmid

Summary The dnaA46 cells can grow at high temperature when a high copy number plasmid pKY31, a derivative of pBR322 carrying a segment of the E. coli chromosome, integrates into the bacterial chromosome. In contrast, the dnaA46 polA ^- cells with the integrated plasmid can not grow at high temperature. Therefore, integration of the plasmid can suppress the dnaA mutation and this suppression requires DNA polymerase I which has been known to be required for plasmid replication. Full reversion of polA or lysogenization of polA ⁺ is lethal for the dnaA46polA ^- bacteria that carry the plasmid only in integrated state. Partial reversion of polA allows these cells to grow at both low and high temperatures. Introduction of the plasmid pBR322 into cytoplasm of these bacteria suppresses the lethal effect caused by full reversion of polA or lysogenization of polA ⁺. This lethal effect expresses independent of the presence or absence of the dnaA mutation. In partial revertants of polA which have only integrated plasmid, the number of copies of a region near the replication origin of integrated plasmid increases. The number is reduced by the presence of extrachromosomal pBR322. It is suggested that the lethal effect of normal levels of DNA polymerase I in strains that carry only the integrated plasmid is due to excessive initiation of replication of the bacterial chromosome from the plasmid origin and high potential of initiation can be absorbed in many copies of cytoplasmic plasmid, probably, in their replication origins.Abbreviations Amp^r ampicillin resistant (resistance) - Tet^s tetracycline sensitive - Tet^r tetracycline resistant - MMS^r methyl methane sulfonate resistant (resistance) - ts temperature sensitive - Kb kilobase pairs     

The tetracycline resistance transposons Tn1721 and Tn1771 have three 38-base-pair repeats and generate five-base-pair direct repeats

Summary The 10.7 kilobase (kb) tetracycline resistance transposons Tn1721 and Tn1771, isolated from disparate sources, are completely homologous on the basis of heteroduplex analyses. Both transposable elements are capable of forming multiple duplications of a 5.3 kb portion encompassing the resistance genes (tet region). A model accounting for both, recA-independent translocation and recA-dependent amplification, postulates two direct and one inverted repeat as essential constituents of the transposons. DNA sequence analyses of Tn1721 and Tn1771 have substantiated this model. They demonstrated three identical 38 base pair repeats identically in both transposons dividing them into a minor transposon and a tet region. Identical sequences of at least 87 base pairs providing recombination hot spots for gene duplication have been found at the ends of the repetitious tet region. Translocation of Tn1721 and Tn1771 generates five base pair direct repeats at the respective sites of insertion. On the basis of the heteroduplex molecules and sequences analyzed the two transposons are identical.To Professor Wolfram Heumann on the occasion of his 65th birthday     

Prevalence of tetracycline resistance genes in Greek seawater habitats

The presence of selected tetracycline resistance (Tc^R) genes was studied in different Greek seawater habitats, originated from wastewater treatment facilities, fishfarm, and coastal environments. The methods employed included assessment of the presence of twelve gene clusters by PCR, followed by hybridization with specific probes, in habitat extracted DNA, Tc^R bacteria, and exogenous isolated plasmids conferring Tc^R. The direct DNA-based analysis showed that tet(A) and tet(K) genes were detected in all habitats, whilst tet(C) and tet(E) were present in fishfarm and wastewater effluent samples and tet(M) was detected in fish-farm and coastal samples. Resistance genes tet(h), tet(C), tet(K), and tet(M) were detected in 60 of the 89 isolates screened. These isolates were identified by fatty acid methyl ester analysis (FAME) as Stenotrophomonas, Acinetobacter, Pseudomonas, Bacillus, and Staphylococcus strains. The presence of the Tc^R genes in 15% of the bacterial isolates coincided with the presence of IncP plasmids. A habitat-specific dissemination of IncP alpha plasmids in wastewater effluent isolates and of IncP beta plasmids in fishfarm isolates was observed. Exogenous isolation demonstrated the presence of plasmids harbouring Tc^R genes in all the habitats tested. Plasmids were shown to carry tet(h), tet(C), tet(E), and tet(K) genes. It is concluded that Tc^R genes are widespread in the seawater habitats studied and often occur on broad host range plasmids that seem to be well disseminated in the bacterial communities.     

Tetracycline Residues and Tetracycline Resistance Genes in Groundwater Impacted by Swine Production Facilities

Antibiotics are used at therapeutic levels to treat disease; at slightly lower levels as prophylactics; and at low, subtherapeutic levels for growth promotion and improvement of feed efficiency. Over 88% of swine producers in the United States gave antimicrobials to grower/finisher pigs in feed as a growth promoter in 2000. It is estimated that ca. 75% of antibiotics are not absorbed by animals and are excreted in urine and feces. The extensive use of antibiotics in swine production has resulted in antibiotic resistance in many intestinal bacteria, which are also excreted in swine feces, resulting in dissemination of resistance genes into the environment.

To assess the impact of manure management on groundwater quality, groundwater samples have been collected near two swine confinement facilities that use lagoons for manure storage and treatment. Several key contaminant indicators—including inorganic ions, antibiotics, and antibiotic resistance genes—were analyzed in groundwater collected from the monitoring wells. Chloride, ammonium, potassium, and sodium were predominant inorganic constituents in the manure samples and served as indicators of groundwater contamination. Based on these analyses, shallow groundwater has been impacted by lagoon seepage at both sites. Liquid chromatography-mass spectroscopy (LC-MS) was used to measure the dissolved concentrations of tetracycline, chlortetracycline, and oxytetracycline in groundwater and manure. Although tetracyclines were regularly used at both facilities, they were infrequently detected in manure samples and then at relatively trace concentrations. Concentrations of all tetracyclines and their breakdown products in the groundwater sampled were generally less than 0.5 μg/L.

Bacterial tetracycline resistance genes served as distinct genotypic markers to indicate the dissemination and mobility of antibiotic resistance genes that originated from the lagoons. Applying PCR to genomic DNA extracted from the lagoon and groundwater samples, four commonly occurring tetracycline (tet) resistance genes—tet(M), tet(O), tet(Q), and tet(W)—were detected. The detection frequency of tet genes was much higher in wells located closer to and down-gradient from the lagoons than in wells more distant from the lagoons. These results suggested that in the groundwater underlying both facilities tetracycline resistance genes exist and are somewhat persistent, but that the distribution and potentially the flux for each tet gene varied throughout the study period.     

Genetic analysis of two bacterial RNA polymerase mutants that inhibit the growth of bacteriophage T7

Summary The Escherichia coli mutants 7009 and BR3 are defective in the growth of bacteriophage T7. We have previously shown that both of these mutant hosts produce an altered RNA polymerase which is resistant to inhibition by the T7 gene 2 protein (De Wyngaert and Hinkle 1979). In both strains, the mutation which prevents T7 growth is closely linked to rifA (rpoB). Both mutants are complemented by transformation with a multicopy plasmid carrying rpoB and rpoC but not by a plasmid carrying only rpoB. This indicates that the mutations reside in rpoC, the structural gene for the subunit of RNA polymerase. When a single copy of the wildtype rpoC allele is introduced into the mutant using the transducing phage drif ^d18, the mutant allele is dominant over wildtype. The drif ^d18 transductant also remains unable to support the growth of T7 in the presence of rifampin. This supports our conclusion that the mutation is in rpoC. We have measured the growth of T7 phage, the kinetics of phage DNA synthesis, and the structure of replicative DNA intermediates in several transductants, and compared these results with those obtained in the original mutant strains.     

Effect of an Insertional Mutation in the cspA Gene Encoding the Major Cold-Shock Protein on Radiation Resistance of Escherichia coli

Plasmid pCspA::Km carrying a cloned mutant allele of the cspA gene for the major Escherichia coli cold-shock protein CspA with an insertion of the kanamycin resistance gene cassette from transposon Tn903 into the core region of the coding sequence causes a 2.3-fold increase in radioresistance of wild-type E. coli cells (cspA ⁺). The radiation protective effect of this plasmid is abolished or drastically reduced in mutants recA13and rpoH15defective in RecA protein and in induction of the heat-shock protein regulon, respectively. Plasmid pCspA::Km causes a 1.3-fold elevation in the resistance to -irradiation of E. coli mutants with an intermediate level of radiation resistance (Gam^r445 and KS0160) but slightly diminishes resistance of a highly radiation-resistant Gam^r444 mutant. In the chromosome of E. coli strain with normal DNA repair systems, the cspA::Km mutation in the homozygous state enhances resistance to the lethal effect of -rays and UV light 2.9 and 1.4 times, respectively. These data suggest that the system of cold-shock proteins can modulate resistance of E. colicells to the lethal effect of -rays and UV light.     

The Escherichia coli proB gene corrects the proline auxotrophy of Saccharomyces cerevisiae pro1 mutants

Summary We constructed plasmids carrying the Escherichia coli proB gene that encodes -glutamyl kinase, under the control of the yeast GAL1 promoter. This construction was carried out with both the wild-type proB ⁺ gene and a mutant allele, proB74, that specifies an enzyme resistant to feedback inhibition by proline. Yeast pro1 mutants harboring these plasmids are proline prototrophs. We conclude that the pro1 mutation results in a deficiency in the -glutamyl kinase activity in Saccharomyces cerevisiae. Expression of the proB74 allele in yeast resulted in enhanced resistance to the proline analogue l-azetidine-2-carboxylate and in a 2.4-fold elevation of the intracellular free proline levels. This result suggests that -glutamyl kinase is the rate limiting step in proline biosynthesis in yeast.     

Molecular analysis of psb A mutations responsible for various herbicide resistance phenotypes in Synechocystis 6714

Mutations conferring herbicide resistance in 3 mutant strains of the cyanobacterium Synechocystis 6714 have been characterized by gene cloning and sequencing. The mutants display very different phenotypes: DCMU-II_A is DCMU-resistant and atrazine-resistant, DCMU-II_B is DCMU-resistant and atrazine-sensitive, and Az-V is DCMU-sensitive, atrazine-resistant and presents particular photoinhibition properties. These mutants were originally obtained either by one-step selection (DCMU-II_A) or by two-step selection (DCMU-II_B and Az-V). psbA copies carrying herbicide resistance have been identified by transformation experiments as psbAI in all cases. Sequences of the psbAI copy of each mutant have been compared to the wild-type sequence. In the single mutant DCMU-II_A, a point mutation at codon 264 (SerAla) results in resistance to both DCMU and atrazine. In the double mutants DCMU-II_B and Az-V, two point mutations were found. DCMU-II_B was derived from DCMU-II_A and had acquired a second mutation at codon 255 (PheLeu) resulting in a slight increase in DCMU resistance and complete abolition of atrazine resistance. Az-V contains two changes at codons 211 (PheSer) and 251 (AlaVal) resulting in high atrazine resistance but only slight DCMU resistance.Abbreviations DCMU: 3-(3,4-dichlorophenyl)-1,1-dimethylurea - PSII: photosystem II     

Recombination deficient mutant of Caulobacter crescentus

Summary A recombination-deficient (Rec^-) strain of Caulobacter crescentus has been isolated from a collection of mutants sensitive to ultraviolet irradiation. The Rec^- mutant fails to give recombinants following Cr30-mediated generalized transduction or following RP4-mediated conjugation. The recombination frequency in the Rec^- strain is at least 5000-fold lower than in the wild type strains. The Rec^- mutant is indistinguishable from wild type in terms of morphology, growth rate, viability, and phage sensitivities, differing only in properties known to be associated with recA-type mutations in other organisms: recombination frequency, ultraviolet sensitivity, and Weigle reactivation. The map location of the rec-526 allele has not been identified, but rec-526 can be cotransferred with the fla-169 mutation by RP4-mediated conjugation at low frequency. This apparent linkage has been used to move the rec mutation to other strains. The Rec^- mutant resembles recA strains of other organisms and provides a healthy strain severely deficient in recombination for use in complementation and cloning studies involving C. crescentus.     

Effect of ogt expression on mutation induction by methyl-, ethyl- and propylmethanesulphonate in Escherichia coli K12 strains

We have previously reported the isolation of an Escherichia coli K12 mutant that is extremely sensitive to mutagenesis by low doses of ethylating agents. We now show by Southern analysis that the mutation involves a gross deletion covering at least the ogt and fnr genes and that no O⁶-alkylguanine-DNA-alkyltransferase activity is present in cell-free extracts of an ada::Tn10 derivative of these bacteria. Confirmation that sensitisation to ethylation-induced mutagenesis was attributable to ogt and not to any other loci covered by the deletion was obtained by constructing derivatives. Thus an ogt::kan^r disruption mutation was introduced into the parental ogt ⁺ bacteria, and the ogt::kan^r mutation was then eliminated by cotransduction of ogt ⁺ with the closely linked Tet ^r marker (zcj::Tn10). The (ogt-fnr) deletion or ogt::kan^r disruption mutants were highly sensitive to ethyl methanesulphonate-induced mutagenesis, as measured by the induction of forward mutations to l-arabinose resistance (Ara¹). Furthermore, the number of Ara^r mutants increased linearly with dose, unlike the case in ogt ⁺ bacteria, which had a threshold dose below which no mutants accumulated. Differences in mutability were even greater with propyl methanesulphonate. Overproduction of the ogt alkyltransferase from a multicopy plasmid reduced ethylmethanesulphonate-induced mutagenesis in the ogt mutant strains and also methylmethanesulphonate mutagenesis in ada ^– bacteria. A sample of AB 1157 obtained from the E. coli K12 genetic stock centre also had a deletion covering the ogt and fnr genes. Since such deletions greatly influence the mutagenic responses to alkylating agents, a survey of the presence of the ogt gene in the E. coli K12 strain being used is advisable.     

Characterization of tetracycline resistance lactobacilli isolated from swine intestines at western area of Taiwan

To investigate the frequency of tetracycline resistance (Tet-R) lactobacilli in pig intestines, a total of 256 pig colons were analyzed and found to contain typical colonies of Tet-R lactic acid bacteria in every sample, ranging from 3.2 × 10³ to 6.6 × 10⁵ CFU/cm². From these samples, a total of 159 isolates of Tet-R lactobacilli were obtained and identified as belonging to 11 species, including Lactobacillus reuteri, Lactobacillus amylovorus, Lactobacillus salivarius, Lactobacillus plantarum, Lactobacillus ruminis, Lactobacillus kefiri, Lactobacillus fermentum, Lactobacillus sakei, Lactobacillus coryniformis, Lactobacillus parabuchneri and Lactobacillus letivazi. Based on the EFSA (2008) breakpoints, all isolates, after MIC analysis, were qualified as Tet-R, from which the significant high Tet-R MIC₅₀ and MIC₉₀ values indicated an ecological distribution of Tet-R lactobacilli mostly with high resistance potency in pig colons. PCR-detection identified 5 tet genes in these isolates, the most predominant one being tet (W), followed by tet (M), (L), (K), and (Q). Their detection rates were 82.0%, 22.5%, 14.4%, 8.1% and 0.9%, respectively. Noteworthily, isolates of the same species carrying identical tet gene(s) usually had a wide different MIC values. Furthermore, strain-subtyping of these isolates by REP-PCR demonstrated a notable genotypic biodiversity % (average = 62%).