scholarly journals Distribution of CRISPR Types in Fluoroquinolone-Resistant Campylobacter jejuni Isolates

Pathogens ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 345
Author(s):  
Mehmet Cemal Adiguzel ◽  
Debora Brito Goulart ◽  
Zuowei Wu ◽  
Jinji Pang ◽  
Seyda Cengiz ◽  
...  
Keyword(s):  
Dna Sequencing ◽  
Campylobacter Jejuni ◽  
Phage Therapy ◽  
Point Mutations ◽  
Common Mutation ◽  
Gyra Gene ◽  
Nucleotide Homology ◽  
Different Sources

To aid development of phage therapy against Campylobacter, we investigated the distribution of the clustered regularly interspaced short palindromic repeats (CRISPR) systems in fluoroquinolone (FQ)-resistant Campylobacter jejuni. A total of 100 FQ-resistant C. jejuni strains from different sources were analyzed by PCR and DNA sequencing to determine resistance-conferring mutation in the gyrA gene and the presence of various CRISPR systems. All but one isolate harbored 1–5 point mutations in gyrA, and the most common mutation was the Thr86Ile change. Ninety-five isolates were positive with the CRISPR PCR, and spacer sequences were found in 86 of them. Among the 292 spacer sequences identified in this study, 204 shared 93–100% nucleotide homology to Campylobacter phage D10, 44 showed 100% homology to Campylobacter phage CP39, and 3 had 100% homology with Campylobacter phage CJIE4-5. The remaining 41 spacer sequences did not match with any phages in the database. Based on the results, it was inferred that the FQ-resistant C. jejuni isolates analyzed in this study were potentially resistant to Campylobacter phages D10, CP39, and CJIE4-5 as well as some unidentified phages. These phages should be excluded from cocktails of phages that may be utilized to treat FQ-resistant Campylobacter.

scholarly journals Accurate mitochondrial DNA sequencing using off-target reads provides a single test to identify pathogenic point mutations

2014 ◽  
Vol 16 (12) ◽  
pp. 962-971 ◽  
Author(s):  
Helen R. Griffin ◽  
Angela Pyle ◽  
Emma L. Blakely ◽  
Charlotte L. Alston ◽  
Jennifer Duff ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Dna Sequencing ◽  
Point Mutations ◽  
Single Test

Prevalence and Antimicrobial Resistance of Campylobacter spp. Isolated from Poultry Carcasses in Poland

2013 ◽  
Vol 76 (8) ◽  
pp. 1451-1455 ◽  
Author(s):  
KINGA WIECZOREK ◽  
IWONA KANIA ◽  
JACEK OSEK
Keyword(s):  
Campylobacter Jejuni ◽  
23S Rrna ◽  
Rrna Gene ◽  
Campylobacter Coli ◽  
Genetic Determinants ◽  
Gyra Gene ◽  
23S Rrna Gene ◽  
Pcr Method ◽  
Almost All ◽  
Campylobacter Spp

The purpose of the present study was to determine the prevalence of Campylobacter in poultry carcasses at slaughter in Poland. For the isolated strains, resistance to selected antibiotics and the associated genetic determinants were identified. A total of 498 Campylobacter isolates were obtained from 802 poultry samples during the 2-year study period. Strains were identified to species with the PCR method; 53.6% of the strains were Campylobacter jejuni and 46.4% were Campylobacter coli. A high percentage of the tested Campylobacter strains were resistant to ciprofloxacin and nalidixic acid (74.1 and 73.5%, respectively) followed by tetracycline (47.4%) and streptomycin (20.5%). Only one C. jejuni and two C. coli isolates were resistant to gentamicin. Seventy-nine (15.9%) of the 498 strains were resistant to three or more classes of antibiotics examined. Higher levels of resistance, irrespective of the antimicrobial agent tested, were found within the C. coli group. Almost all strains resistant to quinolones (99.5%) and to tetracycline (99.6%) carried the Thr-86-to-Ile mutation in the gyrA gene and possessed the tet(O) marker, respectively. All isolates resistant to erythromycin had the A2075G mutation in the 23S rRNA gene. These results reveal that poultry carcasses in Poland are a reservoir of potentially pathogenic and antimicrobial-resistant Campylobacter strains for humans, which may pose a public health risk.

scholarly journals Genomic Heterogeneity and Clonal Evolution in Gastroesophageal Junction Cancer Revealed by Single Cell DNA Sequencing

2021 ◽  
Vol 11 ◽  
Author(s):  
Qingke Duan ◽  
Chao Tang ◽  
Zhao Ma ◽  
Chuangui Chen ◽  
Xiaobin Shang ◽  
...  
Keyword(s):  
Lymph Node ◽  
Lymph Node Metastasis ◽  
Lymph Nodes ◽  
Dna Sequencing ◽  
Single Cell ◽  
Single Cells ◽  
Clonal Evolution ◽  
Gastroesophageal Junction ◽  
Common Mutation ◽  
Node Metastasis

Gastroesophageal junction (GEJ) cancer is a tumor that occurs at the junction of stomach and esophagus anatomically. GEJ cancer frequently metastasizes to lymph nodes, however the heterogeneity and clonal evolution process are unclear. This study is the first of this kind to use single cell DNA sequencing to determine genomic variations and clonal evolution related to lymph node metastasis. Multiple Annealing and Looping Based Amplification Cycles (MALBAC) and bulk exome sequencing were performed to detect single cell copy number variations (CNVs) and single nucleotide variations (SNVs) respectively. Four GEJ cancer patients were enrolled with two (Pt.3, Pt.4) having metastatic lymph nodes. The most common mutation we found happened in the TTN gene, which was reported to be related with the tumor mutation burden in cancers. Significant intra-patient heterogeneity in SNVs and CNVs were found. We identified the SNV subclonal architecture in each tumor. To study the heterogeneity of CNVs, the single cells were sequenced. The number of subclones in the primary tumor was larger than that in lymph nodes, indicating the heterogeneity of primary site was higher. We observed two patterns of multi-station lymph node metastasis: one was skip metastasis and the other was to follow the lymphatic drainage. Taken together, our single cell genomic analysis has revealed the heterogeneity and clonal evolution in GEJ cancer.

scholarly journals Functional Characterization of Excision Repair and RecA-Dependent Recombinational DNA Repair in Campylobacter jejuni

2009 ◽  
Vol 191 (12) ◽  
pp. 3785-3793 ◽  
Author(s):  
Esther J. Gaasbeek ◽  
Fimme J. van der Wal ◽  
Jos P. M. van Putten ◽  
Paulo de Boer ◽  
Linda van der Graaf-van Bloois ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Campylobacter Jejuni ◽  
Uv Irradiation ◽  
Excision Repair ◽  
Spontaneous Mutation ◽  
Functional Characterization ◽  
Point Mutations ◽  
Phase Variable ◽  
Repair Mechanisms

ABSTRACT The presence and functionality of DNA repair mechanisms in Campylobacter jejuni are largely unknown. In silico analysis of the complete translated genome of C. jejuni NCTC 11168 suggests the presence of genes involved in methyl-directed mismatch repair (MMR), nucleotide excision repair, base excision repair (BER), and recombinational repair. To assess the functionality of these putative repair mechanisms in C. jejuni, mutS, uvrB, ung, and recA knockout mutants were constructed and analyzed for their ability to repair spontaneous point mutations, UV irradiation-induced DNA damage, and nicked DNA. Inactivation of the different putative DNA repair genes did not alter the spontaneous mutation frequency. Disruption of the UvrB and RecA orthologues, but not the putative MutS or Ung proteins, resulted in a significant reduction in viability after exposure to UV irradiation. Assays performed with uracil-containing plasmid DNA showed that the putative uracil-DNA glycosylase (Ung) protein, important for initiation of the BER pathway, is also functional in C. jejuni. Inactivation of recA also resulted in a loss of natural transformation. Overall, the data indicate that C. jejuni has multiple functional DNA repair systems that may protect against DNA damage and limit the generation of genetic diversity. On the other hand, the apparent absence of a functional MMR pathway may enhance the frequency of on-and-off switching of phase variable genes typical for C. jejuni and may contribute to the genetic heterogeneity of the C. jejuni population.

Molecular Epidemiology and Resistance Profile of Campylobacter jejuni and Campylobacter coli Strains Isolated from Different Sources in Brazil

2020 ◽  
Vol 26 (12) ◽  
pp. 1516-1525 ◽  
Author(s):  
Carolina N. Gomes ◽  
Miliane R. Frazão ◽  
Jaqueline Passaglia ◽  
Sheila S. Duque ◽  
Marta I. Cazentini Medeiros ◽  
...  
Keyword(s):  
Molecular Epidemiology ◽  
Campylobacter Jejuni ◽  
Campylobacter Coli ◽  
Resistance Profile ◽  
Different Sources

Rapid Detection of Mycobacterium Tuberculosis in Lung Tissue and Analysis of Drug Resistance

2019 ◽  
Vol 11 (5) ◽  
pp. 728-735
Author(s):  
Junlin Chen ◽  
Fei Huang ◽  
Feifan Xu ◽  
Mei Qu ◽  
Delin Gu ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Mycobacterium Tuberculosis ◽  
Dna Sequencing ◽  
Lung Tissue ◽  
Culture System ◽  
Melting Curve ◽  
Gyra Gene ◽  
Tissue Samples ◽  
Curve Method ◽  
Acid Fast Staining

This study investigated the methods for optimizing the workflow for improving the diagnostic efficiency for detection of Mycobacterium tuberculosis (MTB) in lung tissue specimens. A total of 278 specimens were used in this study. M. tuberculosis in fresh lung tissue samples was detected by BACTEC MGIT 960 culture system, culturing L-form MTB, rifampicin (RFP) and levofloxacin (LVFX) susceptibility test, and ribonucleic acid (RNA) simultaneous amplification and testing (SAT). Specimen samples were embedded in paraffin and serially sectioned. The sections were subjected to Ziehl-Neelsen staining and Intensified Kinyoun (IK) acid-fast staining. The suspected MTB or L-form MTB specimens were further investigated by deoxyribonucleic acid (DNA) sequencing and fluorescence polymerase chain reaction (PCR) melting curve method to detect the mutations in rpoB gene and gyrA gene. Thirteen specimens were suspected as MTB positive, and 37 specimens were suspected to be L-form MTB positive by Ziehl-Neelsen staining and IK acid-fast staining. Among the 50 specimens, the number of MTB positive specimens detected by SAT, DNA sequencing, and fluorescence PCR melting curve method was 43, 44, and 45, respectively. Only 11 MTB positive specimens were detected by BACTEC MGIT 960 culture system or by culturing L-form MTB. Mutations detected in rpoB gene and gyrA gene by fluorescence PCR melting curve method were similar to those detected by DNA sequencing. Some specimens, detected by melting curve method, exhibited varied drug resistance to RFP, suggesting heterogeneous resistance. Among the remaining 228 specimens, there was no MTB or L-form MTB detected by BACTEC MGIT 960 culture system. However, 5 specimens were detected to be MTB positive by the SAT method. The fluorescent PCR melting curve method, which has a high level of automation and high sensitivity and specificity, could effectively detect heterozygous drug resistance of MTB in lung tissue samples, which is important for clinicians to effectively formulate a therapeutic strategy.

scholarly journals Bacteriophage Therapy To Reduce Campylobacter jejuni Colonization of Broiler Chickens

2005 ◽  
Vol 71 (11) ◽  
pp. 6554-6563 ◽  
Author(s):  
C. Loc Carrillo ◽  
R. J. Atterbury ◽  
A. El-Shibiny ◽  
P. L. Connerton ◽  
E. Dillon ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Broiler Chickens ◽  
Phage Therapy ◽  
Experimental Models ◽  
Human Infection ◽  
Lytic Bacteriophage ◽  
Bacteriophage Therapy ◽  
Bacteriophage Infection ◽  
Low Passage

ABSTRACT Colonization of broiler chickens by the enteric pathogen Campylobacter jejuni is widespread and difficult to prevent. Bacteriophage therapy is one possible means by which this colonization could be controlled, thus limiting the entry of campylobacters into the human food chain. Prior to evaluating the efficacy of phage therapy, experimental models of Campylobacter colonization of broiler chickens were established by using low-passage C. jejuni isolates HPC5 and GIIC8 from United Kingdom broiler flocks. The screening of 53 lytic bacteriophage isolates against a panel of 50 Campylobacter isolates from broiler chickens and 80 strains isolated after human infection identified two phage candidates with broad host lysis. These phages, CP8 and CP34, were orally administered in antacid suspension, at different dosages, to 25-day-old broiler chickens experimentally colonized with the C. jejuni broiler isolates. Phage treatment of C. jejuni-colonized birds resulted in Campylobacter counts falling between 0.5 and 5 log10 CFU/g of cecal contents compared to untreated controls over a 5-day period postadministration. These reductions were dependent on the phage-Campylobacter combination, the dose of phage applied, and the time elapsed after administration. Campylobacters resistant to bacteriophage infection were recovered from phage-treated chickens at a frequency of <4%. These resistant types were compromised in their ability to colonize experimental chickens and rapidly reverted to a phage-sensitive phenotype in vivo. The selection of appropriate phage and their dose optimization are key elements for the success of phage therapy to reduce campylobacters in broiler chickens.

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