Genetic Rare Variants Affecting Multiple Pathways in Japanese Patients with Palindromic Rheumatism.

Palindromic rheumatism (PR) is a type of cryptogenic paroxysmal arthritis. Several genes may be involved in PR pathogenesis; however, conducting comprehensive case-control genetic studies for PR poses challenges owing to its rarity as a disease. Moreover, case-control studies may overlook rare variants that occur infrequently but play a significant role in pathogenesis. This study aimed to identify disease-related genes in Japanese patients with PR using whole-genome sequencing (WGS) and rare-variant analysis. Genomic DNA was obtained from two familial cases and one sporadic case, and it was subjected to WGS. WGS data of 104 healthy individuals obtained from a public database were used as controls. We performed data analysis for rare variants on detected variants using SKAT-O, KBAC, and SKAT, and subsequently defined significant genes. Significant genes combined with variants shared between the cases were defined as disease-related genes. We also performed pathway analysis for disease-related genes using Reactome. We identified 2,695,244 variants shared between cases; after excluding polymorphisms and noise, 74,640 variants were detected. We identified 540 disease-related genes, including 1,893 variants. Furthermore, we identified 32 significant pathways. Our results indicate that the detected genes and pathways in this study may be involved in PR pathogenesis.

Metagenome and Resistome Analysis of Beta-Lactam-Resistant Bacteria Isolated from River Waters in Surabaya, Indonesia.

Antimicrobial agents are administered to humans and livestock, and bacterial antimicrobial resistance (AMR) and antimicrobial agents are released into the environment. In this study, to investigate the trend of AMR in humans, livestock, and the environment, we performed a metagenomic analysis of multidrug-resistant bacteria with CHROMagar ESBL in environmental river water samples, which were collected using syringe filter units from waters near hospitals, downtown areas, residential areas, and water treatment plants in Surabaya, Indonesia. Our results showed that Acinetobacter, Pseudomonas, Aeromonas, Enterobacter, Escherichia, and Klebsiella grew in CHROMagar ESBL; they were most frequently detected in water samples from rivers surrounding hospitals contaminated with various AMR genes (ARGs) in high levels. These results identified bacteria as ARG reservoirs and revealed that hospitals could be sources for various ARGs disseminated into the environment. In conclusion, this study details a novel metagenomic analysis of collected bacteria in environmental water samples using a syringe filter unit for an AMR epidemiological study based on the One Health approach.

Brothers with Becker muscular dystrophy show discordance in skeletal muscle computed tomography findings: A case report.

Becker muscular dystrophy is caused by DMD mutations and is characterized by progressive muscle atrophy. The wide variations observed in muscle atrophy progression in Becker muscular dystrophy are considered multifactorial, including differences in mutations and environmental factors. In this case, two brothers, aged 2 and 3 years, had the identical DMD mutation, confirming their Becker muscular dystrophy diagnosis. They began using handrails when ascending and descending stairs at the age of 16 due to progressive muscular weakness. Over an 18-year follow-up, the older brother consistently had high serum creatine kinase levels, significantly over median levels. Muscle computed tomography finings revealed that the older brother's gluteus maximus and vastus femoris cross-sectional areas were only half and one-third of the younger brother's, respectively. The mean computed tomography values of gluteus maximus and vastus femoris were significantly lower in the older brother. Our report suggests that muscle atrophy in Becker muscular dystrophy cannot be solely explained by dystrophin mutation or environmental factors.

Association of Neonatal Serum Creatinine Concentration with Maternal Serum Creatinine Concentration and Birth Weight.

BACKGROUND: The serum creatinine (SCr) concentration in neonates is generally high for its body size, compared to those of infants. The aim of the present study was to investigate the effect of maternal SCr on neonatal SCr through measurements of prenatal maternal SCr and neonatal SCr from birth to postnatal Day 5. In addition, postnatal changes in SCr were compared between term and preterm infants, given that few studies have addressed this topic. METHODS: The retrospective study subjects were 151 neonates whose Scr was measured consecutively from birth to postnatal Day 5 and 124 mothers whose SCr was measured prenatally. RESULTS: There were significant correlations between maternal SCr and neonatal SCr at birth (r = 0.858, p < 0.001) and on postnatal Day 1 (r = 0.235, p < 0. 001). The SCr of term infants (median 0.69 mg/dL, range 0.54 - 0.96 mg/ dL) were higher than those of preterm infants (median 0.63 mg/dL, range 0.43 - 1.23 mg/dL, p < 0.001) at birth; however, these values were reversed on postnatal Day 1 (Term: median 0.75 mg/dL, range 0.51 - 1.13 mg/dL, Pre-term: median 0.88 mg/dL, range 0.56 - 1.25 mg/dL, p < 0.001). There were differences in the timing of reaching to peak SCr between preterm and term neonates. In addition, birth weight might affect SCr concentrations after birth. CONCLUSIONS: The results of this study suggest that neonatal SCr is influenced by maternal SCr, although the effect disappears by postnatal Day 2. Moreover, glomerular filtration rate differs between term and preterm infants.

Molecular characteristics and genetic analysis of the genes related to carbapenemase, efflux pump, and outer membrane proteins in carbapenem-resistant Klebsiella pneumoniae.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is a life-threatening pathogen that has not been fully investigated on a molecular basis. Therefore, the molecular mechanisms of carbapenem resistance in CRKP collected from medical institutions in Hyogo Prefecture has been analyzed. Antimicrobial susceptibilities and the presence of carbapenemase along with epidemiological analyzes using multilocus sequence typing (MLST) have been investigated. The relative expression of efflux pump genes and mutations of ompK35 and ompK36, encoding the outer membrane porin, were also assessed for their relationship with carbapenem resistance. Most of the collected 22 CRKP isolates were non-susceptible to imipenem (68.2%), meropenem (90.9%), and ertapenem (81.8%), but all 22 strains were susceptible to colistin. Twelve strains (54.5%) were detected for carbapenemase genes such as blaIMP-6. Sequence type 37 was detected by MLST in 10 strains (45.5%). Non-carbapenemase-producing strains had high resistance rates for three carbapenems, and the main cause of resistance was ompK35 mutation. In conclusion, the main cause of resistance was imipenemase metallo-ß-lactamase (IMP-6) production in carbapenemase-producing strains, and ompK35 mutation in non-carbapenemase-producing strains. Susceptibility to carbapenem did not differ in CRKP regardless of carbapenemase production, except for imipenem susceptibility. This result contributes to a more insightful understanding of the mechanisms of CRKP in Japan.

Comparative genetic analysis of the antimicrobial susceptibilities and virulence of hypermucoviscous and non-hypermucoviscous ESBL-producing Klebsiella pneumoniae in Japan.

BACKGROUND: Hypermucoviscous (HMV) Klebsiella pneumoniae produces large amounts of capsular polysaccharides, leading to high mortality. Since extended spectrum beta-lactamase (ESBL)-producing HMV K. pneumoniae strains have increased in Japan, we investigated and compared the antimicrobial susceptibilities and genetic characteristics of HMV and non-HMV ESBL-producing K. pneumoniae. METHODS: We investigated 291 ESBL-producing K. pneumoniae collected between 2012 and 2018, and in them 54 HMV strains were identified and comparable 53 non-HMV strains were selected. Then, ESBL gene detection, plasmid replicon typing, and virulence gene detection were done by PCR amplification. RESULTS: Almost all of the HMV K. pneumoniae strains possessed uge (98.1%), wabG (96.3%), rmpA (94.4%), iucA (79.6%), fimH (70.4%), iroB (70.4%), and peg-344 (70.4%). These genes were found less frequently in non-HMV strains (uge 20.8%, wabG 83.0%, rmpA 7.5%, iucA 3.8%, fimH 9.4%, iroB 5.7%, and peg-344 1.9%). K2 capsule type (40.7%) was most common in HMV strains. HMV strains showed higher resistance to cefepime (p = 0.001) and piperacillin/tazobactam (p = 0.005) than non-HMV strains. CTX-M-15 (75.9%, 60.4%) was the dominant ESBL type in both HMV and non-HMV strains, and the most common plasmid replicon type was IncFII (52.1%) in CTX-M-15-producing strains. CONCLUSIONS: We found that HMV strains had more virulence genes and showed higher resistance to antibiotics than non-HMV strains. The most common capsule type was K2. CTX-M-15 was the most common type of ESBL gene in both HMV and non-HMV strains in Japan. The FII plasmid might be related to the spread of CTX-M-15 among K. pneumoniae strains.

Impact on quinolone resistance of plasmid-mediated quinolone resistance gene and mutations in quinolone resistance-determining regions in extended spectrum beta lactamase-producing Klebsiella pneumoniae isolated from urinary tract infection patients.

Klebsiella pneumoniae is a typical pathogen in urinary tract infections (UTI), and the emergence of extended spectrum beta-lactamase (ESBL)-producing strains has been frequently reported, accompanied by higher quinolone resistance rates. There are two major mechanisms of quinolone resistance, mutations in quinolone resistance-determining regions (QRDR) and the presence of the plasmid-mediated quinolone resistance (PMQR) genes. This study aimed to investigate quinolone resistance among 105 ESBL-producing K. pneumoniae specimens isolated from UTI patients in Indonesia. These were characterized for antimicrobial resistance to nalidixic acid, ciprofloxacin, and levofloxacin, QRDR mutations in gyrA and parC and the presence of PMQR genes. We found that 84.8% of the collected isolates were resistant to at least one of the quinolones. QRDR mutation in gyrA was observed in 49.5% of these strains and parC mutations in 61.0%. PMQR genes were identified in 84.8% of strains. The QRDR mutations clearly had a greater effect on resistance than the PMQR genes. In conclusion, we found high quinolone resistance rates in Indonesian ESBL-producing K. pneumoniae, in which QRDR mutation played a major role.

Genetic characteristics of azithromycin-resistant Neisseria gonorrhoeae collected in Hyogo, Japan during 2015-2019.

Introduction. Azithromycin (AZM) is a therapeutic drug for sexually transmitted infections and is used for Neisseria gonorrhoeae when first- and second-line drugs are not available. Recently, the susceptibility of N. gonorrhoeae against AZM has been decreasing worldwide.Hypothesis/Gap Statement. Azithromycin-resistance (AZM-R) rates among N. gonorrhoeae in Japan are increasing, and the gene mutations and epidemiological characteristics of AZM-R in N. gonorrhoeae have not been fully investigated.Aim. We determined the susceptibility to AZM and its correlation with genetic characteristics of N. gonorrhoeae.Methodology. We investigated the susceptibility to AZM and genetic characteristics of N. gonorrhoeae. Mutations in domain V of the 23S rRNA gene and mtrR were examined in 93 isolates, including 13 AZM-R isolates. Spread and clonality were examined using sequence types (STs) of multi-antigen sequence typing for N. gonorrhoeae (NG-MAST), and whole genome analysis (WGA) to identify single nucleotide polymorphisms.Results. The number of AZM-R isolates increased gradually from 2015 to 2019 in Hyogo (P=0.008). C2599T mutations in 23S rRNA significantly increased in AZM-R isolates (P<0.001). NG-MAST ST4207 and ST6762 were frequently detected in AZM-R isolates, and they had higher MICs to AZM from 6 to 24 µg/ml. The phylogenic tree-based WGA showed that all isolates with ST4207 were contained in the same clade, and isolates with ST6762 were divided into two clades, AZM-S isolates and AZM-R isolates, which were different from the cluster containing ST1407.Conclusion. Our study showed yearly increases in AZM-R rates in N. gonorrhoeae. NG-MAST ST4207 and ST6762 were not detected in our previous study in 2015 and were frequently identified in isolates with higher MICs to AZM. WGA confirmed that isolates with these STs are closely related to each other. Continued surveillance is needed to detect the emergence and confirm the spread of NG-MAST ST4207 and ST6762.

Antibiotic Resistance in Non-Typhoidal Salmonella enterica Strains Isolated from Chicken Meat in Indonesia.

The increase in antibiotic resistance in non-typhoidal Salmonella enterica (NTS) has been confirmed in Indonesia by this study. We confirmed the virulence genes and antimicrobial susceptibilities of clinical NTS (n = 50) isolated from chicken meat in Indonesia and also detected antimicrobial resistance genes. Of 50 strains, 30 (60%) were non-susceptible to nalidixic acid (NA) and all of them had amino acid mutations in gyrA. Among 27 tetracycline (TC) non-susceptible strains, 22 (81.5%) had tetA and/or tetB. The non-susceptibility rates to ampicillin, gentamicin or kanamycin were lower than that of NA or TC, but the prevalence of blaTEM or aadA was high. Non-susceptible strains showed a high prevalence of virulence genes compared with the susceptible strains (tcfA, p = 0.014; cdtB, p < 0.001; sfbA, p < 0.001; fimA, p = 0.002). S. Schwarzengrund was the most prevalent serotype (23 strains, 46%) and the most frequently detected as multi-antimicrobial resistant. The prevalence of virulence genes in S. Schwarzengrund was significantly higher than other serotypes in hlyE (p = 0.011) and phoP/Q (p = 0.011) in addition to the genes above. In conclusion, NTS strains isolated from Indonesian chicken had a high resistance to antibiotics and many virulence factors. In particular, S. Schwarzengrund strains were most frequently detected as multi-antimicrobial resistant and had a high prevalence of virulence genes.

A sandwich ELISA kit reveals marked elevation of titin N-terminal fragment levels in the urine of mdx mice.

The mdx mouse is a model of Duchenne muscular dystrophy (DMD), a fatal progressive muscle wasting disease caused by dystrophin deficiency, and is used most widely in preclinical studies. Mice with dystrophin deficiency, however, show milder muscle strength phenotypes than humans. In human, the introduction of a sandwich enzyme-linked immunosorbent assay (ELISA) kit revealed a more than 700-fold increase in titin N-terminal fragment levels in the urine of pediatric patients with DMD. Notably, the urinary titin level declines with aging, reflecting progression of muscle wasting. In mouse, development of a highly sensitive ELISA kit has been awaited. Here, a sandwich ELISA kit to measure titin N-terminal fragment levels in mouse urine was developed. The developed kit showed good linearity, recovery, and repeatability in measuring recombinant or natural mouse titin N-terminal fragment levels. The titin N-terminal fragment concentration in the urine of mdx mice was more than 500-fold higher than that of normal mice. Urinary titin was further analyzed by extending the collection of urine samples to both young (3-11 weeks old) and aged (56-58 weeks old) mdx mice. The concentration in the young group was significantly higher than that in the aged group. It was concluded that muscle protein breakdown is active and persistent in mdx mice even though the muscle phenotype is mild. Our results provide an opportunity to develop DMD treatments that aim to alleviate muscle protein breakdown by monitoring urinary titin levels.

Occurrence of Carriage of Multidrug Resistant Enterobacteriaceae among Pregnant Women in the Primary Health Center and Hospital Setting in Surabaya, Indonesia.

Objectives: The incidence of healthy individuals carrying multidrug resistant Enterobacteriaceae, including extended-spectrum ß-lactamase producing Enterobacteriaceae (ESBL-E), especially extended-spectrum ß-lactamase producing Escherichia coli (ESBL-EC) and extended-spectrum ß-lactamase producing Klebsiella pneumoniae (ESBL-KP), is increasing worldwide. Although ESBL-E causes early or late onset of neonatal sepsis, the prevalence of ESBL-E carriage among pregnant women in Indonesia is not clear. In the present study, we compared the occurrence of carriage of ESBL-E among pregnant women in a primary health center (PHC) versus two hospitals. Materials and Methods: We collected rectal swab samples from 200 pregnant women who visited a PHC or were admitted to two hospitals in Surabaya, Indonesia from July to October 2018. The ESBL-E strains were isolated from the samples and phenotypically and genotypically analyzed. Results: ESBL-E strains were isolated from 25 (24.8%) pregnant women who visited the PHC and 49 (49.5%) pregnant women who were admitted to the hospitals. The rate of ESBL-E carriage of pregnant women in the hospitals was significantly higher than that in the PHC. Among the 74 isolated ESBL-E strains, ESBL-EC was most frequently isolated (62 strains), followed by ESBL-KP (12 strains). In addition, blaCTX-M-15 was the most frequent ESBL gene type of the isolated ESBL-E strains. Conclusions: Our results revealed the high occurrence of ESBL-E carriage in pregnant women, especially those who were admitted to the hospitals.

Assessment of catabolic state in infants with the use of urinary titin N-fragment.

BACKGROUND: Urinary titin N-fragment levels have been used to assess the catabolic state, and we used this biomarker to evaluate the catabolic state of infants. METHODS: We retrospectively measured urinary titin N-fragment levels of urinary samples. The primary outcome was its changes according to postmenstrual age. The secondary outcomes included differences between gestational age, longitudinal change after birth, influence on growth, and relationship with blood tests. RESULTS: This study included 219 patients with 414 measurements. Urinary titin N-fragment exponentially declined with postmenstrual age. These values were 12.5 (7.1-19.6), 8.1 (5.1-13.0), 12.8 (6.0-21.3), 26.4 (16.4-52.0), and 81.9 (63.3-106.4) pmol/mg creatinine in full, late, moderate, very, and extremely preterm infants, respectively (p < 0.01). After birth, urinary levels of titin N-fragment exponentially declined, and the maximum level within a week was associated with the time to return to birth weight in preterm infants (ρ = 0.39, p < 0.01). This was correlated with creatine kinase in full-term infants (ρ = 0.58, p < 0.01) and with blood urea nitrogen in preterm infants (ρ = 0.50, p < 0.01). CONCLUSIONS: The catabolic state was increased during the early course of the postmenstrual age and early preterm infants. IMPACT: Catabolic state in infants, especially in preterm infants, was expected to be increased, but no study has clearly verified this. In this retrospective study of 219 patients with 414 urinary titin measurements, the catabolic state was exponentially elevated during the early postmenstrual age. The use of the urinary titin N-fragment clarified catabolic state was prominently increased in very and extremely preterm infants.

Antibacterial Activity of Membrane-Permeabilizing Bactericidal Cyclodextrin Derivatives.

Antimicrobial peptides that act by disrupting bacterial membranes are attractive agents for treating drug-resistant bacteria. This study investigates a membrane-disrupting peptide mimic made of a cyclic oligosaccharide cyclodextrin scaffold that can be chemically polyfunctionalized. An antibacterial functional group on the peptide was simplified to an alkylamino group that combines cationic and hydrophobic moieties, the former to interact with the anionic bacterial membrane and the latter with the membrane interior. The cyclodextrins equipped with eight alkylamino groups on the molecules using a poly-click reaction exhibited antibacterial activity against Gram-positive and Gram-negative bacteria, including drug-resistant pathogens such as carbapenem-resistant Enterobacteriaceae. Several lines of evidence showed that these agents disrupt bacterial membranes, leading to rapid bacterial cell death. The resulting membrane perturbation was directly visualized using high-speed atomic force microscopy imaging. In Gram-negative bacteria, the membrane-permeabilizing action of these derivatives allowed the entry of co-treated traditional antibiotics, which were then active against these bacteria.

Effect of Antimicrobial Stewardship on Oral Quinolone Use and Resistance Patterns over 8 Years (2013-2020).

Since 2014, several global and national guidelines have been introduced to address the problem of antimicrobial resistance. We conducted a campaign in a tertiary hospital to promote appropriate quinolone use through educational lectures in 2018. The aim of this retrospective study was to evaluate the changes in the following: prescription characteristics, trend of oral quinolone use, and antibiotic susceptibility of bacteria from 2013 to 2020. Antimicrobial use was assessed as days of therapy per 1000 patient-days. We found a significant reduction in unnecessary antibiotic prescriptions between December 2013 and December 2020. Significant negative trends were detected in the use of quinolones over 8 years (outpatients, coefficient = -0.15655, p < 0.001; inpatients, coefficient = -0.004825, p = 0.0016). In particular, the monthly mean use of quinolones among outpatients significantly decreased by 11% from 2013 to 2014 (p < 0.05) and reduced further by 31% from 2017 to 2020 (p < 0.001). A significant positive trend was observed in the susceptibility of Pseudomonas aeruginosa to levofloxacin (p < 0.001). These results demonstrate that the use of oral quinolones was further reduced following educational intervention and the bacterial susceptibility improved with optimal quinolone usage compared to that in 2013.

Impact of Cefazolin Shortage on Clinical Outcomes of Adult Patients with Bacteremia Caused by Methicillin-Susceptible Staphylococcus aureus in a Tertiary Care University Hospital.

Cefazolin is an essential antibiotic used for treating bacteremia; in particular, it is recommended as a first-line agent for infections caused by methicillin-susceptible Staphylococcusaureus (MSSA). In March 2019, problems with a major antibiotic supplier caused a critical shortage of cefazolin in Japan; however, the impact of the cefazolin shortage on clinical outcomes remains unknown. This study aimed to evaluate the effect of the cefazolin shortage in patients with MSSA bacteremia. Data from 75 patients were compared between the pre-shortage (March 2018-January 2019, n = 39) and post-shortage (March 2019-January 2020, n = 36) periods. There were no significant differences in the demographic characteristics between the two groups, and the cefazolin shortage did not worsen clinical outcomes such as adverse drug reactions, treatment failure, and 30-day mortality. In the post-shortage group, ampicillin/sulbactam and benzylpenicillin were more frequently administered as alternative antibiotics for empirical and definitive therapy (10% vs. 31%, p = 0.042; 0% vs. 19%, p = 0.004, respectively). Multivariate analysis revealed that the broad-spectrum antibiotics for definitive therapy, such as antipseudomonal penicillin, were associated with treatment failure in patients with MSSA bacteremia (OR = 17, p = 0.003). Hence, narrow-spectrum antibiotics should be prescribed for MSSA bacteremia as alternatives during a cefazolin shortage.

Differential effects of chromosome and plasmid blaCTX-M-15 genes on antibiotic susceptibilities in extended-spectrum beta-lactamase-producing Escherichia coli isolates from patients with urinary tract infection.

OBJECTIVES: To compare antibiotic susceptibilities between chromosomal and plasmid blaCTX-M-15 locations in urinary tract infection-causing extended-spectrum ß-lactamases-producing Escherichia coli blaCTX-M-15 isolated in Indonesia. METHODS: A total of 84 strains identified as extended-spectrum ß-lactamases-producing E. coli were isolated from patients with urinary tract infection in Indonesia in 2015. Antimicrobial susceptibility tests were performed on these strains using 18 antibiotics, and extended-spectrum ß-lactamase bla genes were detected by polymerase chain reaction. Gene localization of blaCTX-M-15 -positive strains was confirmed by Southern blot hybridization, and epidemiological typing was conducted using multilocus sequence typing. RESULTS: Of 54 strains harboring the blaCTX-M-15 gene, 27 showed localization on chromosome, 20 on plasmid, and seven on chromosome and plasmid. Most multilocus sequence typing sequence types of the 27 strains with chromosomal blaCTX-M-15 were ST405 (25.9%) and ST131 (22.2%) strains, whereas the 20 strains with plasmid-blaCTX-M-15 were mostly ST410 (55.0%). CONCLUSIONS: Extended-spectrum ß-lactamases-producing E. coli blaCTX-M-15 with plasmid genes show significantly higher resistant rates against piperacillin-tazobactam but lower resistant rates against chloramphenicol compared to chromosomal strains in Indonesian patients with urinary tract infection. Mechanistic investigations will be necessary to advance our knowledge of antimicrobial resistance in urinary tract infection.

Cross-Resistance and the Mechanisms of Cephalosporin-Resistant Bacteria in Urinary Tract Infections Isolated in Indonesia.

Urinary tract infection (UTI) by antibiotic-resistant strains has become increasingly problematic, with trends that differ from country to country. This study examined cross-resistance and the mechanisms of cephalosporin resistance in UTI-causative bacteria isolated in Indonesia. Antibiotic susceptibility tests based on Clinical Laboratory Standards Institute (CLSI) standards were done for UTI-causative strains (n = 50) isolated from patients in Indonesia in 2015-2016 and showed resistance against the third-generation cephalosporin. Mechanistic studies were carried out to confirm the presence of extended-spectrum ß-lactamase (ESBL) genes, carbapenemase-related genes, the fosA3 gene related to fosfomycin resistance, and mutations of quinolone-resistance-related genes. Isolated UTI-causative bacteria included Escherichia coli (64.0%), Pseudomonas aeruginosa (16.0%), Klebsiella pneumoniae (10.0%), and others (10.0%). These strains showed 96.0% susceptibility to amikacin, 76.0% to fosfomycin, 90.0% to imipenem, 28.0% to levofloxacin, 92.0% to meropenem, and 74.0% to tazobactam/piperacillin. ESBL was produced by 68.0% of these strains. Mechanistic studies found no strains with carbapenemase genes but 6.0% of strains had the fosA3 gene. Seventy-two % of the strains had mutations in the gyrA gene and 74.0% in the parC gene. Most E. coli strains (87.5%) had Ser-83 → Leu and Asp-87 → Asn in gyrA and 93.8% of E. coli had Ser-80 → Ile in parC. There were significant correlations among mutations in gyrA and parC, and fosA3 gene detection (P < 0.05), respectively. To our knowledge, this is the first mechanistic study of antibiotic-cross-resistant UTI-causative bacteria in Indonesia. Further studies with a longer period of observation are necessary, especially for changes in carbapenem resistance without carbapenemase-related genes.

Risk Factors for the Acquisition of Enterococcus faecium Infection and Mortality in Patients with Enterococcal Bacteremia: A 5-Year Retrospective Analysis in a Tertiary Care University Hospital.

The incidence of bacteremia caused by Enterococcus faecium, which is highly resistant to multiple antibiotics, is increasing in Japan. However, risk factors for the acquisition of E. faecium infection and mortality due to enterococcal bacteremia are not well known. We compared demographic, microbiological, and clinical characteristics using a Cox regression model and univariate analysis. We performed a multivariate analysis to identify risk factors for patients treated between 2014 and 2018. Among 186 patients with enterococcal bacteremia, two groups included in the Kaplan-Meier analysis (E. faecalis (n = 88) and E. faecium (n = 94)) showed poor overall survival in the E. faecium group (HR: 1.92; 95% confidence interval: 1.01-3.66; p = 0.048). The median daily antibiotic cost per patient in the E. faecium group was significantly higher than that in the E. faecalis group ($23 ($13-$34) vs. $34 ($22-$58), p < 0.001). E. faecium strains were more frequently identified with previous use of antipseudomonal penicillins (OR = 4.04, p < 0.001) and carbapenems (OR = 3.33, p = 0.003). Bacteremia from an unknown source (OR = 2.79, p = 0.025) and acute kidney injury (OR = 4.51, p = 0.004) were associated with higher risks of 30-day mortality in patients with enterococcal bacteremia. Therefore, clinicians should provide improved medical management, with support from specialized teams such as those assisting antimicrobial stewardship programs.

Genetic analysis of ESBL-producing Klebsiella pneumoniae isolated from UTI patients in Indonesia.

INTRODUCTION: Extended spectrum beta-lactamase (ESBL)-producing Klebsiellapneumoniae is a serious concern for nosocomial infection and the emergence rate in Indonesia is higher than that in developed countries. The purpose of this study was to investigate the genetic characteristics of ESBL-producing K. pneumoniae isolated from UTI patients in Indonesia. MATERIALS AND METHODS: We collected K. pneumoniae resistant to ceftazidime or cefotaxime isolated from UTI patients in Dr. Soetomo's Academic Hospital in Surabaya, Indonesia in 2015. Ninety-four strains were identified as ESBL-producing bacteria by confirmation tests. The isolates were investigated by antimicrobial susceptibility testing with 20 drugs and ESBL gene detection, plasmid replicon typing and virulence genes as hypermucoviscous (HMV) strains were tested by the string test. RESULTS: High rates of resistance to ciprofloxacin (86.2%), tetracycline (80.9%) and nalidixic acid (78.7%) were observed. CTX-M-15 was the most common ESBL gene (89.4%), 33 of which also carried SHV-type ESBL. IncF was the most prevalent plasmid replicon typing (47.6%). Sixteen (17.0%) strains were judged as HMV, all of which had rmpA and more than half of which had fimH, uge, and wab. IncL/M was the most common replicon plasmid in the HMV strains, and the difference in the positive rate was statistically significant (p = 0.0024). CONCLUSION: This study showed the high prevalence of multiple-drug resistant and predominately CTX-M-15-positive ESBL-producing K. pneumoniae in Indonesia. There was a correlation between IncL/M and the HMV phenotype in this study. As such hypervirulent strains continue to emerge, studying their dissemination with resistance determinants is an urgent priority.

The Antimicrobial Resistance Characteristics of Imipenem-Non-Susceptible, Imipenemase-6-Producing Escherichia coli.

Imipenemase-6 (IMP-6) type carbapenemase-producing Enterobacteriaceae is regarded as dangerous due to its unique lack of antimicrobial susceptibility. It is resistant to meropenem (MEPM) but susceptible to imipenem (IPM). In addition to carbapenemase, outer membrane porins and efflux pumps also play roles in carbapenem resistance by reducing the antimicrobial concentration inside cells. Extended-spectrum ß-lactamase (ESBL) is transmitted with IMP-6 by the plasmid and broadens the spectrum of antimicrobial resistance. We collected 42 strains of IMP-6-producing Escherichia coli and conducted a molecular analysis of carbapenemase, ESBL, porin, efflux, and epidemiological characteristics using plasmid replicon typing. Among the 42 isolates, 21 strains were susceptible to IPM (50.0%) and 1 (2.4%) to MEPM. Seventeen strains (40.5%) co-produced CTX-M-2 type ESBL. We found that the relative expression of ompC and ompF significantly correlated with the MIC of IPM (p = 0.01 and p = 0.03, respectively). Sixty-eight% of CTX-M-2-non-producing strains had IncI1, which was significantly different from CTX-M-2-producing strains (p < 0.001). In conclusion, 50.0% of our IMP-6-producing strains were non-susceptible to IPM, which is different from the typical pattern and can be attributed to decreased porin expression. Further studies investigating other types of carbapenemase are warranted.