Constitutive and Inducible Expression of the rRNA Methylase Gene erm(B) in Campylobacter

Macrolides are the antimicrobials of choice for treating human campylobacteriosis. The recent emergence of erm(B) in Campylobacter bacteria threatens the utility of this class of antibiotics. Here we report the constitutive and inducible expression of erm(B) in Campylobacter isolates derived from diarrheal patients and food-producing animals. Constitutive expression of erm(B) was associated with insertion and deletion in the regulatory region of the gene, providing the first documentation of the differential expression of erm(B) in Campylobacter bacteria

Characterization of photosystem II in transgenic tobacco plants with decreased iron superoxide dismutase

AbstractIron superoxide dismutases (FeSODs) play an important role in preventing the oxidative damage associated with photosynthesis. To investigate the mechanisms of FeSOD in protection against photooxidative stress, we obtained transgenic tobacco (Nicotiana tabacum) plants with severely decreased FeSOD by using a gene encoding tobacco chloroplastic FeSOD for the RNAi construct. Transgenic plants were highly sensitive to photooxidative stress and accumulated increased levels of O2•− under normal light conditions. Spectroscopic analysis and electron transport measurements showed that PSII activity was significantly reduced in transgenic plants. Flash-induced fluorescence relaxation and thermoluminescence measurements revealed that there was a slow electron transfer between QA and QB and decreased redox potential of QB in transgenic plants, whereas the donor side function of PSII was not affected. Immunoblot and blue native gel analyses showed that PSII protein accumulation was also decreased in transgenic plants. PSII photodamage and D1 protein degradation under high light treatment was increased in transgenic plants, whereas the PSII repair was not affected, indicating that the stability of the PSII complex was decreased in transgenic plants. The results in this study suggest that FeSOD plays an important role in maintaining PSII function by stabilizing PSII complexes in tobacco plants

Investigation of Haemophilus parasuis from healthy pigs in China

Haemophilus parasuis is a common colonizer of the upper respiratory tract of swine and frequently causes disease, especially in weaner pigs. To date, limited epidemiological data was available for H. parasuis from healthy pigs, which might be carriers of potential pathogenic strains. In this study, from September 2016 to October 2017, we investigated the prevalence and characteristics of H. parasuis from healthy pigs in China. Totally, we obtained 244 isolates from 1675 nasal samples from 6 provinces. H. parasuis isolation was more successful in weaner pigs (22.6%, 192/849), followed by finisher pigs (9.3%, 43/463), and sows (2.5%, 9/363). The most prevalent serovars were 7 (20.1%, 49/244), followed by 3 (14.8%, 36/244), 2 (14.3%, 35/244), 11 (12.7%, 31/244), 5/12 (5.7%, 14/244) and 4 (2.5%, 6/244). Bimodal or multimodal distributions of MICs were observed for most of the tested drugs, which suggested the presence of non-wild type populations. It was noted that the MIC90 values of tilmicosin (64 μg/ml) was relatively higher than that reported in previous studies. Our results suggest that: 1) potentially pathogenic serovars of H. parasuis are identified in healthy pigs, and 2) elevated MICs and presence of mechanisms of resistance not yet described for clinically important antimicrobial agents would increase the burden of disease caused by H. parasuis.info:eu-repo/semantics/acceptedVersio

Emergence of Multidrug-Resistant Campylobacter Species Isolates with a Horizontally Acquired rRNA Methylase

Antibiotic-resistant Campylobacter constitutes a serious threat to public health, and resistance to macrolides is of particular concern, as this class of antibiotics is the drug of choice for clinical therapy of campylobacteriosis. Very recently, a horizontally transferrable macrolide resistance mediated by the rRNA methylase gene erm(B) was reported in a Campylobacter coli isolate, but little is known about the dissemination of erm(B) among Campylobacter isolates and the association of erm(B)-carrying isolates with clinical disease. To address this question and facilitate the control of antibiotic-resistant Campylobacter, we determined the distribution of erm(B) in 1,554 C. coli and Campylobacter jejuni isolates derived from food-producing animals and clinically confirmed human diarrheal cases. The results revealed that 58 of the examined isolates harbored erm(B) and exhibited high-level resistance to macrolides, and most were recent isolates, derived in 2011-2012. In addition, the erm(B)-positive isolates were all resistant to fluoroquinolones, another clinically important antibiotic used for treating campylobacteriosis. The erm(B) gene is found to be associated with chromosomal multidrug resistance genomic islands (MDRGIs) of Gram-positive origin or with plasmids of various sizes. All MDRGIs were transferrable to macrolide-susceptible C. jejuni by natural transformation under laboratory conditions. Molecular typing of the erm(B)-carrying isolates by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) identified diverse genotypes and outbreak-associated diarrheal isolates. Molecular typing also suggested zoonotic transmission of erm(B)-positive Campylobacter. These findings reveal an emerging and alarming trend of dissemination of erm(B) and MDRGIs in Campylobacter and underscore the need for heightened efforts to control their further spread

Identification of a Novel Genomic Island Conferring Resistance to Multiple Aminoglycoside Antibiotics in Campylobacter coli

Historically, the incidence of gentamicin resistance in Campylobacter has been very low, but recent studies reported a high prevalence of gentamicin-resistant Campylobacter isolated from food-producing animals in China. The reason for the high prevalence was unknown and was addressed in this study. PCR screening identified aminoglycoside resistance genes aphA-3 and aphA-7 and the aadE–sat4–aphA-3 cluster among 41 Campylobacter isolates from broiler chickens. Importantly, a novel genomic island carrying multiple aminoglycoside resistance genes was identified in 26 aminoglycoside resistant Campylobacter coli strains. Sequence analysis revealed that the genomic island was inserted between cadF and COO1582 on the C. coli chromosome and consists of 14 open reading frames (ORFs), including 6 genes (the aadE–sat4–aphA-3 cluster, aacA-aphD, aac, and aadE) encoding aminoglycoside-modifying enzymes. Analysis by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing indicated that the C. coli isolates carrying this unique genomic island were clonal, and the clone of PFGE subtype III and sequence type (ST) 1625 was particularly predominant among the C. coli isolates examined, suggesting that clonal expansion may be involved in dissemination of this resistance island. Additionally, we were able to transfer this genomic island from C. coli to a Campylobacter jejuni strain using natural transformation under laboratory conditions, and the transfer resulted in a drastic increase in aminoglycoside resistance in the recipient strain. These findings identify a previously undescribed genomic island that confers resistance to multiple aminoglycoside antibiotics. Since aminoglycoside antibiotics are used for treating occasional systemic infections caused by Campylobacter, the emergence and spread of this antibiotic resistance genomic island represent a potential concern for public health

Transferable Multiresistance Plasmids Carrying cfr in Enterococcus spp. from Swine and Farm Environment

Seventy-seven porcine Enterococcus isolates with florfenicol MICs of ≥16 μg of were/ml screened for the presence of the multiresistance gene cfr, its location on plasmids, and its genetic environment. Three isolates—Enterococcus thailandicus 3-38 (from a porcine rectal swab collected at a pig farm), Enterococcus thailandicus W3, and Enterococcus faecalis W9-2 (the latter two from sewage at a different farm), carried the cfr gene. The SmaI pulsed-field gel electrophoresis patterns of the three isolates differed distinctly. In addition, E. faecalis W9-2 was assigned to a new multilocus sequence type ST469. Mating experiments and Southern blot analysis indicated that cfr is located on conjugative plasmids pW3 (∼75 kb) from E. thailandicus W3, p3-38 (∼72 kb) from E. thailandicus 3-38, and pW9-2 (∼55 kb) from E. faecalis W9-2; these plasmids differed in their sizes, additional resistance genes, and the analysis of the segments encompassing the cfr gene. Sequence analysis revealed that all plasmids harbored a 4,447-bp central region, in which cfr was bracketed by two copies of the novel insertion sequence ISEnfa4 located in the same orientation. The sequences flanking the central regions of these plasmids, including the partial tra gene regions and a ω-ε-ζ toxin-antitoxin module, exhibited \u3e95% nucleotide sequence identity to the conjugative plasmid pAMβ1 from E. faecalis. Conjugative plasmids carrying cfr appear to play an important role in the dissemination and maintenance of the multiresistance gene cfr among enterococcal isolates and possibly other species of Gram-positive bacteria

Distribution of the Multidrug Resistance Gene cfr in Staphylococcus Species Isolates from Swine Farms in China

A total of 149 porcine Staphylococcus isolates with florfenicol MICs of ≥16 μg/ml were screened for the presence of the multiresistance gene cfr, its location on plasmids, and its genetic environment. In total, 125 isolates carried either cfr (16 isolates), fexA (92 isolates), or both genes (17 isolates). The 33 cfr-carrying staphylococci, which included isolates of the species Staphylococcus cohnii, S. arlettae, and S. saprophyticus in which the cfr gene has not been described before, exhibited a wide variety of SmaI pulsed-field gel electrophoresis patterns. In 18 cases, the cfr gene was located on plasmids. Four different types of cfr-carrying plasmids—pSS-01 (n = 2; 40 kb), pSS-02 (n = 3; 35.4 kb), pSS-03 (n = 10; 7.1 kb), and pBS-01 (n = 3; 16.4 kb)—were differentiated on the basis of their sizes, restriction patterns, and additional resistance genes. Sequence analysis revealed that in plasmid pSS-01, the cfr gene was flanked in the upstream part by a complete aacA-aphD-carrying Tn4001-like transposon and in the downstream part by a complete fexA-carrying transposon Tn558. In plasmid pSS-02, an insertion sequence IS21-558 and the cfr gene were integrated into transposon Tn558 and thereby truncated the tnpA and tnpB genes. The smallest cfr-carrying plasmid pSS-03 carried the macrolide-lincosamide-streptogramin B resistance gene erm(C). Plasmid pBS-01, previously described in Bacillus spp., harbored a Tn917-like transposon, including the macrolide-lincosamide-streptogramin B resistance gene erm(B) in the cfr downstream region. Plasmids, which in part carry additional resistance genes, seem to play an important role in the dissemination of the gene cfr among porcine staphylococci

Identification of a Novel G2073A Mutation in 23S rRNA in Amphenicol-Selected Mutants of Campylobacter jejuni

Objectives This study was conducted to examine the development and molecular mechanisms of amphenicol resistance in Campylobacter jejuni by using in vitro selection with chloramphenicol and florfenicol. The impact of the resistance development on growth rates was also determined using in vitro culture. Methods Chloramphenicol and florfenicol were used as selection agents to perform in vitro stepwise selection. Mutants resistant to the selective agents were obtained from the selection process. The mutant strains were compared with the parent strain for changes in MICs and growth rates. The 23S rRNA gene and the L4 and L22 ribosomal protein genes in the mutant strains and the parent strain were amplified and sequenced to identify potential resistance-associated mutations. Results C. jejuni strains that were highly resistant to chloramphenicol and florfenicol were obtained from in vitro selection. A novel G2073A mutation in all three copies of the 23S rRNA gene was identified in all the resistant mutants examined, which showed resistance to both chloramphenicol and florfenicol. In addition, all the mutants selected by chloramphenicol also exhibited the G74D modification in ribosomal protein L4, which was previously shown to confer a low-level erythromycin resistance in Campylobacter species. The mutants selected by florfenicol did not have the G74D mutation in L4. Notably, the amphenicol-resistant mutants also exhibited reduced susceptibility to erythromycin, suggesting that the selection resulted in cross resistance to macrolides. Conclusions This study identifies a novel point mutation (G2073A) in 23S rRNA in amphenicol-selected mutants of C. jejuni. Development of amphenicol resistance in Campylobacter likely incurs a fitness cost as the mutant strains showed slower growth rates in antibiotic-free media