Effect of passive versus active abdominal drainage on wound infection after pancreatectomy: A meta-analysis.

Following pancreatic resection, there may be a variety of complications, including wound infection, haemorrhage, and abdominal infection. The placement of drainage channels during operation may decrease the chances of postoperative complications. However, what kind of drainage can decrease the rate of postoperative complications is still a matter of debate. The purpose of this research is to evaluate the efficacy of both active and passive drainage for post-operation wound complications. From the beginning of the database until November 2023, EMBASE, the Cochrane Library and the Pubmed database have been searched. The two authors collected 2524 related studies from 3 data bases for importation into Endnote software, and 8 finished trials were screened against the exclusion criteria. Passive drainage can decrease the incidence of superficial wound infection in postoperative patients with pancreas operation (Odds Ratio [OR], 1.30; 95% CI, 1.06-1.60 p = 0.01); No statistically significant difference was found in the incidence of deep infections among the two groups (OR, 1.51; 95% CI, 0.68-3.36 p = 0.31); No statistical significance was found for the rate of haemorrhage after active drainage on the pancreas compared with that of passive drainage (OR, 0.72; 95% CI, 0.29-1.77 p = 0.47); No statistically significant difference was found in the rate of death after operation for patients who had received a pancreas operation in active or passive drainage (OR, 0.90; 95% CI, 0.57-1.42 p = 0.65); On the basis of existing evidence, the use of passive abdominal drainage reduces postoperative surface wound infections in patients. But there were no statistically significant differences in the risk of severe complications, haemorrhage after surgery, or mortality. However, because of the limited sample size of this meta-analysis, it is necessary to have more high-quality research with a large sample size to confirm the findings.

Single-minded homolog 2 as a potential prognostic signature and assessment of its correlation with immune cell infiltration in pancreatic cancer.

Single-minded homolog 2 (SIM2) has been identified as a potential contributor to the development of solid tumors. Despite this, there is a lack of comprehensive research regarding its biological role and underlying mechanism within pancreatic cancer (PC), as well as its prognostic impact. This study systematically evaluated the expression level and clinical significance of SIM2 in patients with PC using various databases, including The Cancer Genome Atlas, KM Plotter, and gene expression profiling interactive analysis. To investigate the relationship between SIM2 expression and immune cell infiltration, we conducted ESTIMATE and single-sample gene set enrichment analysis (ssGSEA) analyses. Single-minded homolog 2 was up-regulated in patients with PC. Pancreatic cancer patients with higher SIM2 expression had poorer overall survival rates. Gene set enrichment analysis results suggested that SIM2 may have a significant impact on the progression of PC and the regulation of immune responses. According to the ssGSEA algorithm, SIM2 has a negative correlation with the levels of infiltrating TFH, mast cells, and pDC. Our study demonstrated that SIM2 serves as a biomarker, and is associated with both prognosis and immune infiltration in PC. This provides a solid foundation for future investigations into the precise role of SIM2 in the carcinogenesis and progression of PC.

The Modulating Mechanisms of miRNA-196 in Malignancies and Its Prognostic Value: A Systematic Review and Meta-Analysis.

Accumulating studies have revealed that up- or downregulated miRNA-196 expression correlates with the prognostic value in various malignancies; however, existing single studies lack robust evidence to elucidate the role of miRNA-196 in malignancy. The pooled results showed that the upregulation of miRNA-196 expression was significantly correlated with unfavorable OS [HR 2.14; 95% confidence interval (CI), 1.78-2.57; p < 0.001)] and worse PFS (HR 2.84; 95% CI, 1.29-6.23, P = 0.01) in various malignancies. According to the regulatory mechanisms, studies shown that multiple tumors associated with transcription processes could be modulated by the miRNA-196 family; correspondingly, the miRNA-196 family exerted biological functions that could be regulated by various molecules. The upregulation of miRNA-196a, miRNA-196b and miRNA-196 expression is correlated with significantly unfavorable OS in multiple malignancies; similarly, miRNA-196 overexpression predicts poor PFS in multiple malignancies. Taken together, these findings indicate that miRNA-196a and miRNA-196b may serve as oncogenic molecules and may be potential prognostic biomarkers in multiple malignancies.

Prognostic Nomogram for patients undergoing radical Pancreaticoduodenectomy for adenocarcinoma of the pancreatic head.

BACKGROUND: Radical pancreaticoduodenectomy is the most common treatment strategy for patients diagnosed with adenocarcinoma of the pancreatic head. Few studies have reported the clinical characteristics and treatment efficacies of patients undergoing radical pancreaticoduodenectomy for adenocarcinoma of the pancreatic head. METHODS: A total of 177 pancreatic head cancer patients who underwent radical pancreaticoduodenectomy and were pathologically confirmed as having pancreatic ductal adenocarcinoma were screened in the West China Hospital of Sichuan University. The multivariate analysis results were implemented to construct a nomogram. The concordance index (c-index), the area under the curve (AUC) and calibration were utilized to evaluate the predictive performance of the nomogram. RESULTS: The prognostic nutritional index (PNI), the lymph node ratio (LNR) and the American Joint Committee on Cancer (AJCC) staging served as independent prognostic factors and were used to construct the nomogram. The c-indexes of the nomogram were 0.799 (confidence interval (CI), 0.741-0.858) and 0.732 (0.657-0.807) in the primary set and validation set, respectively. The AUCs of the nomogram at 1 and 3 years were 0.832 and 0.783, which were superior to the AJCC staging values of 0.759 and 0.705, respectively. CONCLUSIONS: The nomogram may be used to predict the prognosis of radical resection for adenocarcinoma of the pancreatic head. These findings may represent an effective model for the developing an optimal therapeutic schedule for malnourished patients who need early effective nutritional intervention and may promote the treatment efficacy of resectable adenocarcinoma of the pancreatic head.

Five gene signatures were identified in the prediction of overall survival in resectable pancreatic cancer.

BACKGROUND: Although genes have been previously detected in pancreatic cancer (PC), aberrant genes that play roles in resectable pancreatic cancer should be further assessed. METHODS: Messenger RNA samples and clinicopathological data corrected with PC were downloaded from The Cancer Genome Atlas (TCGA). Resectable PC patients were randomly divided into a primary set and a validation set. Univariable Cox regression analysis, lasso-penalized Cox regression analysis, and multivariable Cox analysis were implemented to distinguish survival-related genes (SRGs). A risk score based on the SRGs was calculated by univariable Cox regression analysis. A genomic-clinical nomogram was established by integrating the risk score and clinicopathological data to predict overall survival (OS) in resectable PC. RESULTS: Five survival-related genes (AADAC, DEF8, HIST1H1C, MET, and CHFR) were significantly correlated with OS in resectable PC. The resectable PC patients, based on risk score, were sorted into a high-risk group that showed considerably unfavorable OS (p < 0.001) than the low-risk group, in both the primary set and the validation set. The concordance index (C-index) was calculated to evaluate the predictive performance of the nomogram were respectively in the primary set [0.696 (0.608-0.784)] and the validation set [0.682 (0.606-0.758)]. Additionally, gene set enrichment Analysis discovered several meaningful enriched pathways. CONCLUSION: Our study identified five prognostic gene biomarkers for OS prediction and which facilitate postoperative molecular target therapy for the resectable PC, especially the nomic-clinical nomogram which may be used as an effective model for the postoperative OS evaluation and also an optimal therapeutic tool for the resectable PC.

The Rho-Independent Transcription Terminator for the porA Gene Enhances Expression of the Major Outer Membrane Protein and Campylobacter jejuni Virulence in Abortion Induction.

Campylobacter jejuni is a leading cause of foodborne illnesses worldwide. Its porA gene encodes the major outer membrane protein (MOMP) that is abundantly expressed and has important physiological functions, including a key role in systemic infection and abortion induction in pregnant animals. Despite the importance of porA in C. jejuni pathogenesis, mechanisms modulating its expression levels remain elusive. At the 3' end of the porA transcript, there is a Rho-independent transcription terminator (named T porA in this study). Whether T porA affects the expression and function of MOMP remains unknown and is investigated in this study. Green fluorescent protein (GFP) fusion constructs with the porA promoter at the 5' end and an intact T porA or no T porA at the 3' end of the gfp coding sequence revealed that both the transcript level of gfp and its fluorescence signals were more than 2-fold higher in the construct with T porA than in the one without T porA Real-time quantitative PCR (qRT-PCR) analysis of the porA mRNA and immunoblot detection of MOMP in C. jejuni showed that disruption of T porA significantly reduced the porA transcript level and the expression of MOMP. An mRNA decay assay demonstrated that disruption of T porA resulted in a shortened transcript half-life of the upstream gfp or porA gene, indicating that T porA enhances mRNA stability. In the guinea pig model, the C. jejuni construct with an interrupted T porA was significantly attenuated in abortion induction. Together, these results indicate that T porA enhances the expression level of MOMP by stabilizing its mRNA and influences the virulence of C. jejuni.

Risk Factors for Carbapenem-Resistant Pseudomonas aeruginosa, Zhejiang Province, China.

Carbapenem-resistant Pseudomonas aeruginosa (CRPA) is a public health concern worldwide, but comprehensive analysis of risk factors for CRPA remains limited in China. We conducted a retrospective observational study of carbapenem resistance in 71,880 P. aeruginosa isolates collected in Zhejiang Province during 2015-2017. We analyzed risk factors for CRPA, including the type of clinical specimen; the year, season, and region in which it was collected; patient information, including age, whether they were an outpatient or inpatient, and whether inpatients were in the intensive care unit or general ward; and the level of hospital submitting isolates. We found CRPA was more prevalent among isolates from patients >60 years of age and in inpatients, especially in intensive care units. In addition, specimen types and seasons in which they were collected were associated with higher rates of CRPA. Our findings can help hospitals reduce the spread of P. aeruginosa and optimize antimicrobial drug use.

Characterization of multiresistance gene cfr(C) variants in Campylobacter from China.

OBJECTIVES: To investigate the occurrence, the genetic environment and the functionality of novel variants of the MDR gene cfr(C) in Campylobacter from China. METHODS: A total of 370 Campylobacter isolates of porcine and chicken origin collected from three regions of China in 2015 were screened for cfr(C) by PCR. The phenotypes and genotypes of cfr(C)-positive isolates were investigated by antimicrobial susceptibility testing, PFGE, MLST, S1-PFGE, Southern blotting and WGS. Quantitative RT-PCR was used to compare the expression levels of the cfr(C) variants in their original isolate and clone constructs in Campylobacter jejuni NCTC 11168. RESULTS: Four (1.1%) porcine Campylobacter coli isolates were positive for cfr(C). They failed to show elevated MICs of phenicols. The deduced Cfr(C) sequences identified exhibited 2-6 amino acid changes compared with the original Cfr(C) reported in the USA. Cloning of the cfr(C) variant genes into C. jejuni NCTC 11168 resulted in ≥32-fold increases in the MICs of phenicols, indicating that the cfr(C) variant genes are functional. The cfr(C)-carrying isolates belonged to three genotypes and WGS analysis revealed the cfr(C) genes were chromosomally located in MDR genomic islands, which contained multiple antibiotic resistance genes of Gram-positive origin. CONCLUSIONS: This study identified chromosomal cfr(C) genes in C. coli isolates from China. They appeared functionally dormant in the original isolates but were fully functional when cloned and expressed in C. jejuni. The cfr(C) genes were co-transferred with other antibiotic resistance genes, possibly from Gram-positive bacteria. These findings reveal new insights into the function and transmission of cfr(C) in Campylobacter.

High Prevalence of Fluoroquinolone-Resistant Campylobacter Bacteria in Sheep and Increased Campylobacter Counts in the Bile and Gallbladders of Sheep Medicated with Tetracycline in Feed.

Campylobacter is a major foodborne pathogen in humans and a significant cause of abortion in sheep. Although ruminants are increasingly recognized as important reservoirs for Campylobacter species, limited information is available about the molecular epidemiology and antimicrobial resistance (AMR) profiles of sheep Campylobacter Here, we describe a two-trial study that examined Campylobacter profiles in sheep and determined whether in-feed tetracycline (TET) influenced the distribution and AMR profiles of Campylobacter Each trial involved 80 commercial sheep naturally infected with Campylobacter: 40 of these sheep were medicated with tetracycline in feed, while the other 40 received feed without antibiotics. Fecal and bile samples were collected for the isolation of Campylobacter The bacterial isolates were analyzed for antimicrobial susceptibility and genotypes. The results revealed that 87.0% and 61.3% of the fecal and bile samples were positive for Campylobacter (Campylobacter jejuni and Campylobacter coli), with no significant differences between the medicated and nonmedicated groups. All but one of the tested Campylobacter isolates were resistant to tetracycline. Although fluoroquinolone (FQ) resistance remained low in C. jejuni (1.7%), 95.0% of the C. coli isolates were resistant to FQ. Genotyping revealed that C. jejuni sequence type 2862 (ST2862) and C. coli ST902 were the predominant genotypes in the sheep. Feed medication with tetracycline did not affect the overall prevalence, species distribution, and AMR profiles of Campylobacter, but it did increase the total Campylobacter counts in bile and gallbladder. These findings identify predominant Campylobacter clones, reveal the high prevalence of FQ-resistant C. coli, and provide new insights into the epidemiology of Campylobacter in sheep.IMPORTANCECampylobacter is a major cause of foodborne illness in humans, and antibiotic-resistant Campylobacter is considered a serious threat to public health in the United States and worldwide. As a foodborne pathogen, Campylobacter commonly exists in the intestinal tract of ruminant animals, such as sheep and cattle. Results from this study reveal the predominant genotypes and high prevalence of tetracycline (TET) and fluoroquinolone (FQ) resistance in sheep Campylobacter The finding on fluoroquinolone resistance in sheep Campylobacter is unexpected, as this class of antibiotics is not used for sheep in the United States, and it may suggest the transmission of fluoroquinolone-resistant Campylobacter from cattle to sheep. Additionally, the results demonstrate that in-feed medication with tetracycline increases Campylobacter counts in gallbladders, suggesting that the antibiotic promotes Campylobacter colonization of the gallbladder. These findings provide new information on Campylobacter epidemiology in sheep, which may be useful for curbing the spread of antibiotic-resistant Campylobacter in animal reservoirs.

Investigating the Suitability of a Laboratory Mouse Model to Study the Pathogenesis of Abortifacient Campylobacter jejuni.

The aim of this study was to assess whether pregnant mice represent a useful model to study the reproductive pathology of Campylobacter jejuni IA3902 using the end point of positive microbial culture of the organism from the fetoplacental unit. Pregnant BALB/c and CD-1 mice (14 days' gestation) were inoculated orally and intraperitoneally (IP) with 1 × 109 colony-forming units/ml of C. jejuni IA3902. The organism was recovered by microbial culture from the fetoplacental unit in 10 of 10 BALB/c and 10 of 10 CD-1 IP-inoculated pregnant mice and in 29% (2/7) BALB/c and 38% (3/8) CD-1 orally inoculated pregnant mice. Gross reproductive lesions included necrosuppurative placentitis, fetal resorption, intrauterine fetal death, stillborn pups (dead neonates), and multifocal hepatitis. Histological changes consisted of locally extensive neutrophilic and necrotizing placentitis with intralesional bacterial colonies of C. jejuni, ulcerative endometritis, random multifocal hepatitis, and rare cholecystitis. Immunohistochemistry for the major outer membrane protein of C. jejuni revealed moderate to large numbers of the organism at the periphery of the placental discs, within trophoblasts and extracellularly, with invasion into the placental disc largely via the vascular network. The organism is trophic for neutral mucin, iron, and L-fucose within the murine placenta. C. jejuni IA3902 has affinity for the murine reproductive tract, specifically the fetoplacental unit, where it results in a necrotizing placentitis with positive microbial recovery after both IP and oral challenge in BALB/c and CD-1 pregnant mice.

Point mutations in the major outer membrane protein drive hypervirulence of a rapidly expanding clone of Campylobacter jejuni.

Infections due to clonal expansion of highly virulent bacterial strains are clear and present threats to human and animal health. Association of genetic changes with disease is now a routine, but identification of causative mutations that enable disease remains difficult. Campylobacter jejuni is an important zoonotic pathogen transmitted to humans mainly via the foodborne route. C. jejuni typically colonizes the gut, but a hypervirulent and rapidly expanding clone of C. jejuni recently emerged, which is able to translocate across the intestinal tract, causing systemic infection and abortion in pregnant animals. The genetic basis responsible for this hypervirulence is unknown. Here, we developed a strategy, termed "directed genome evolution," by using hybridization between abortifacient and nonabortifacient strains followed by selection in an animal disease model and whole-genome sequence analysis. This strategy successfully identified SNPs in porA, encoding the major outer membrane protein, are responsible for the hypervirulence. Defined mutagenesis verified that these mutations were both necessary and sufficient for causing abortion. Furthermore, sequence analysis identified porA as the gene with the top genome-wide signal of adaptive evolution using Fu's Fs, a population genetic metric for recent population size changes, which is consistent with the recent expansion of clone "sheep abortion." These results identify a key virulence factor in Campylobacter and a potential target for the control of this zoonotic pathogen. Furthermore, this study provides general, unbiased experimental and computational approaches that are broadly applicable for efficient elucidation of disease-causing mutations in bacterial pathogens.

Genotypes and Antimicrobial Susceptibility Profiles of Hemolytic Escherichia coli from Diarrheic Piglets.

Hemolytic Escherichia coli are important pathogens in neonatal and weaned pigs. In this study, we analyzed 95 hemolytic E. coli isolated from intestinal contents or fecal samples of diarrheic piglets in 15 states of the United States between November 2013 and December 2014. Phenotypic antimicrobial susceptibility was determined through Sensititre BOFO6F plates for all the strains. They were all resistant to clindamycin, penicillin, tiamulin, tilmicosin, and highly resistant to oxytetracycline (91.6%), chlortetracycline (78.9%), ampicillin (75.8%), and sulfadimethoxine (68.4%). 86.2% of them were multidrug resistant. Whole genome sequencing (WGS) showed that 55 strains were enterotoxigenic E. coli (ETEC) and 40 strains were non-ETEC, and the strains belonged to 22 known and 2 novel sequence types (STs). ST100 and ST10 were the main and predominant STs in ETEC strains, whereas the non-ETEC strains were diverse with ST23 and ST761 as the main STs. Antibiotic resistance gene/mutation profiling of the genomes confirmed the results of antimicrobial susceptibility test. Notably, significant differences were found in the susceptibility to enrofloxacin between ETEC and non-ETEC (58.2% vs. 5.0%) and gentamicin (32.7% vs. 7.5%). ampH, ampC2, and ampC1 were the most common beta-lactamase genes in all E. coli strains, and extended-spectrum beta-lactamase (ESBL) genes were rare in these isolates. This study provides new insights into antibiotic resistance and genotypes of intestinal pathogenic E. coli associated with swine disease in the United States, and support the utility of WGS in accurate prediction of resistance to most antibiotics.

High Genetic Plasticity in Multidrug-Resistant Sequence Type 3-IncHI2 Plasmids Revealed by Sequence Comparison and Phylogenetic Analysis.

We report a novel fusion plasmid, pP2-3T, cointegrating sequence type 3 (ST3)-IncHI2 with an IncFII plasmid backbone mediating multidrug resistance (MDR) and virulence. Phylogenetic analysis and comparative genomics revealed that pP2-3T and other MDR ST3-IncHI2 plasmids clustered together, representing a unique IncHI2 lineage that exhibited high conservation in backbones of plasmids but possessed highly genetic plasticity in various regions by acquiring numerous antibiotic resistance genes and fusing with other plasmids. Surveillance studies should be performed to monitor multiresistance IncHI2 plasmids among Enterobacteriaceae.

A Cotransformation Method To Identify a Restriction-Modification Enzyme That Reduces Conjugation Efficiency in Campylobacter jejuni.

Conjugation is an important mechanism for horizontal gene transfer in Campylobacter jejuni, the leading cause of human bacterial gastroenteritis in developed countries. However, to date, the factors that significantly influence conjugation efficiency in Campylobacter spp. are still largely unknown. Given that multiple recombinant loci could independently occur within one recipient cell during natural transformation, the genetic materials from a high-frequency conjugation (HFC) C. jejuni strain may be cotransformed with a selection marker into a low-frequency conjugation (LFC) recipient strain, creating new HFC transformants suitable for the identification of conjugation factors using a comparative genomics approach. To test this, an erythromycin resistance selection marker was created in an HFC C. jejuni strain; subsequently, the DNA of this strain was naturally transformed into NCTC 11168, an LFC C. jejuni strain, leading to the isolation of NCTC 11168-derived HFC transformants. Whole-genome sequencing analysis and subsequent site-directed mutagenesis identified Cj1051c, a putative restriction-modification enzyme (aka CjeI) that could drastically reduce the conjugation efficiency of NCTC 11168 (>5,000-fold). Chromosomal complementation of three diverse HFC C. jejuni strains with CjeI also led to a dramatic reduction in conjugation efficiency (∼1,000-fold). The purified recombinant CjeI could effectively digest the Escherichia coli-derived shuttle vector pRY107. The endonuclease activity of CjeI was abolished upon short heat shock treatment at 50°C, which is consistent with our previous observation that heat shock enhanced conjugation efficiency in C. jejuni Together, in this study, we successfully developed and utilized a unique cotransformation strategy to identify a restriction-modification enzyme that significantly influences conjugation efficiency in C. jejuniIMPORTANCE Conjugation is an important horizontal gene transfer mechanism contributing to the evolution of bacterial pathogenesis and antimicrobial resistance. Campylobacter jejuni, the leading foodborne bacterial organism, displays significant strain diversity due to horizontal gene transfer; however, the molecular components influencing conjugation efficiency in C. jejuni are still largely unknown. In this study, we developed a cotransformation strategy for comparative genomics analysis and successfully identified a restriction-modification enzyme that significantly influences conjugation efficiency in C. jejuni The new cotransformation strategy developed in this study is also expected to be broadly applied in other naturally competent bacteria for functional comparative genomics research.

Key Role of Capsular Polysaccharide in the Induction of Systemic Infection and Abortion by Hypervirulent Campylobacter jejuni.

Campylobacter jejuni is a zoonotic pathogen, and a hypervirulent clone, named clone SA, has recently emerged as the predominant cause of ovine abortion in the United States. To induce abortion, orally ingested Campylobacter must translocate across the intestinal epithelium, spread systemically in the circulation, and reach the fetoplacental tissue. Bacterial factors involved in these steps are not well understood. C. jejuni is known to produce capsular polysaccharide (CPS), but the specific role that CPS plays in systemic infection and particularly abortion in animals remains to be determined. In this study, we evaluated the role of CPS in bacteremia using a mouse model and in abortion using a pregnant guinea pig model following oral challenge. Compared with C. jejuni NCTC 11168 and 81-176, a clone SA isolate (IA3902) resulted in significantly higher bacterial counts and a significantly longer duration of bacteremia in mice. The loss of capsule production via gene-specific mutagenesis in IA3902 led to the complete abolishment of bacteremia in mice and abortion in pregnant guinea pigs, while complementation of capsule expression almost fully restored these phenotypes. The capsule mutant strain was also impaired for survival in guinea pig sera and sheep blood. Sequence-based analyses revealed that clone SA possesses a unique CPS locus with a mosaic structure, which has been stably maintained in all clone SA isolates derived from various hosts and times. These findings establish CPS as a key virulence factor for the induction of systemic infection and abortion in pregnant animals and provide a viable candidate for the development of vaccines against hypervirulent C. jejuni.

Identification and functional analysis of two toxin-antitoxin systems in Campylobacter jejuni.

Toxin-antitoxin (TA) systems are widely distributed in bacteria and play an important role in maintaining plasmid stability. The leading foodborne pathogen, Campylobacter jejuni, can carry multiple plasmids associated with antibiotic resistance or virulence. Previously a virulence plasmid named pVir was identified in C. jejuni 81-176 and IA3902, but determining the role of pVir in pathogenesis has been hampered because the plasmid cannot be cured. In this study, we report the identification of two TA systems that are located on the pVir plasmid in 81-176 and IA3902, respectively. The virA (proteic antitoxin)/virT (proteic toxin) pair in IA3902 belongs to a Type II TA system, while the cjrA (RNA antitoxin)/cjpT (proteic toxin) pair in 81-176 belongs to a Type I TA system. Notably, cjrA (antitoxin) represents the first noncoding small RNA demonstrated to play a functional role in Campylobacter physiology to date. By inactivating the TA systems, pVir was readily cured from Campylobacter, indicating their functionality in Campylobacter. Using pVir-cured IA3902, we demonstrated that pVir is not required for abortion induction in the guinea pig model. These findings establish the key role of the TA systems in maintaining plasmid stability and provide a means to evaluate the function of pVir in Campylobacter pathobiology.

Emergence of a plasmid-borne multidrug resistance gene cfr(C) in foodborne pathogen Campylobacter.

Objectives: To identify and characterize a novel cfr variant that recently emerged and confers multidrug resistance in Campylobacter , a major foodborne pathogen. Methods: WGS was initially used to identify the cfr (C) gene in Campylobacter isolates and its function was further verified by cloning into an antibiotic-susceptible Campylobacter jejuni strain. Distribution of cfr (C) in various Campylobacter isolates was determined by PCR analysis. Genotyping of cfr (C)-positive strains was done by PFGE and MLST. Results: The cfr (C) gene is predicted to encode a protein that shares 55.1% and 54.9% identity with Cfr and Cfr(B), respectively. cfr (C) was located on a conjugative plasmid of ∼48 kb. Cloning of cfr (C) into C. jejuni NCTC 11168 and conjugative transfer of the cfr (C)-containing plasmid confirmed its role in conferring resistance to phenicols, lincosamides, pleuromutilins and oxazolidinones, and resulted in an 8-256-fold increase in their MICs in both C. jejuni and Campylobacter coli . The cfr (C) gene was detected in multiple C. coli (34 of 344; 10%) isolates derived from different cattle farms in different states, and molecular typing of the cfr (C)-positive C. coli isolates revealed its spread mainly via clonal expansion. Conclusions: These results identify cfr (C) as a new multidrug resistance mechanism in Campylobacter and suggest the potential transmission of this mechanism via the foodborne route, warranting enhanced efforts to monitor its spread in Campylobacter and other foodborne pathogens.

A zero-inflated Poisson model for insertion tolerance analysis of genes based on Tn-seq data.

MOTIVATION: Transposon insertion sequencing (Tn-seq) is an emerging technology that combines transposon mutagenesis with next-generation sequencing technologies for the identification of genes related to bacterial survival. The resulting data from Tn-seq experiments consist of sequence reads mapped to millions of potential transposon insertion sites and a large portion of insertion sites have zero mapped reads. Novel statistical method for Tn-seq data analysis is needed to infer functions of genes on bacterial growth. RESULTS: In this article, we propose a zero-inflated Poisson model for analyzing the Tn-seq data that are high-dimensional and with an excess of zeros. Maximum likelihood estimates of model parameters are obtained using an expectation-maximization (EM) algorithm, and pseudogenes are utilized to construct appropriate statistical tests for the transposon insertion tolerance of normal genes of interest. We propose a multiple testing procedure that categorizes genes into each of the three states, hypo-tolerant, tolerant and hyper-tolerant, while controlling the false discovery rate. We evaluate the proposed method with simulation studies and apply the proposed method to a real Tn-seq data from an experiment that studied the bacterial pathogen, Campylobacter jejuniAvailability and implementation: We provide R code for implementing our proposed method at http://github.com/ffliu/TnSeq A user's guide with example data analysis is also available there. CONTACT: pliu@iastate.edu SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Wide but Variable Distribution of a Hypervirulent Campylobacter jejuni Clone in Beef and Dairy Cattle in the United States.

Campylobacter jejuni clone SA is the major cause of sheep abortion and contributes significantly to foodborne illnesses in the United States. Clone SA is hypervirulent because of its distinct ability to produce systemic infection and its predominant role in clinical sheep abortion. Despite the importance of clone SA, little is known about its distribution and epidemiological features in cattle. Here we describe a prospective study on C. jejuni clone SA prevalence in 35 feedlots in 5 different states in the United States and a retrospective analysis of clone SA in C. jejuni isolates collected by National Animal Health Monitoring System (NAHMS) dairy studies in 2002, 2007, and 2014. In feedlot cattle feces, the overall prevalence of Campylobacter organisms was 72.2%, 82.1% of which were C. jejuni Clone SA accounted for 5.8% of the total C. jejuni isolates, but its prevalence varied by feedlot and state. Interestingly, starlings on the feedlots harbored C. jejuni in feces, including clone SA, suggesting that these birds may play a role in the transmission of Campylobacter In dairy cattle, the overall prevalence of clone SA was 7.2%, but a significant decrease in the prevalence was observed from 2002 to 2014. Whole-genome sequence analysis of the dairy clone SA isolates revealed that it was genetically stable over the years and most of the isolates carried the tetracycline resistance gene tet(O) in the chromosome. These findings indicate that clone SA is widely distributed in both beef and dairy cattle and provide new insights into the molecular epidemiology of clone SA in ruminants.IMPORTANCEC. jejuni clone SA is a major cause of small-ruminant abortion and an emerging threat to food safety because of its association with foodborne outbreaks. Cattle appear to serve as a major reservoir for this pathogenic organism, but there is a major gap in our knowledge about the epidemiology of clone SA in beef and dairy cattle. By taking advantage of surveillance studies conducted on a national scale, we found a wide but variable distribution of clone SA in feedlot cattle and dairy cows in the United States. Additionally, the work revealed important genomic features of clone SA isolates from cattle. These findings provide critically needed information for the development of preharvest interventions to control the transmission of this zoonotic pathogen. Control of C. jejuni clone SA will benefit both animal health and public health, as it is a zoonotic pathogen causing disease in both ruminants and humans.

A single nucleotide change in mutY increases the emergence of antibiotic-resistant Campylobacter jejuni mutants.

OBJECTIVES: Mutator strains play an important role in the emergence of antibiotic-resistant bacteria. Campylobacter jejuni is a leading cause of foodborne illnesses worldwide and is increasingly resistant to clinically important antibiotics. The objective of this study was to identify the genetic basis that contributes to a mutator phenotype in Campylobacter and determine the role of this phenotype in the development of antibiotic resistance. METHODS: A C. jejuni isolate (named CMT) showing a mutator phenotype was subjected to WGS analysis. Comparative genomics, site-specific reversion and mutation, and gene knockout were conducted to prove the mutator effect was caused by a single nucleotide change in the mutY gene of C. jejuni. RESULTS: The C. jejuni CMT isolate showed ∼ 100-fold higher mutation frequency to ciprofloxacin than the WT strain. Under selection by ciprofloxacin, fluoroquinolone-resistant mutants emerged readily from the CMT isolate. WGS identified a single nucleotide change (G595 → T) in the mutY gene of the CMT isolate. Further experiments using defined mutant constructs proved its specific role in elevating mutation frequencies. The mutY point mutation also led to an ∼ 700-fold increase in the emergence of ampicillin-resistant mutants, indicating its broader impact on antibiotic resistance. Structural modelling suggested the G595 → T mutation probably affects the catalytic domain of MutY and consequently abolishes the anti-mutator function of this DNA repair protein. CONCLUSIONS: The G595 → T mutation in mutY abolishes its anti-mutator function and confers a mutator phenotype in Campylobacter, promoting the emergence of antibiotic-resistant Campylobacter.