High body mass index and triglyceride levels at health checkups increase the risk of new-onset chronic kidney disease and worsening renal function: the TAMA MED Project-CKD.

BACKGROUND: Health checkups are important in patients with chronic kidney disease (CKD), which is not easily accompanied by subjective symptoms. CKD can be caused or aggravated by factors that have not yet been identified. METHODS: This retrospective cohort study included 7 483 patients who underwent specific annual health checkups at a medical institution in Tama City, did not have CKD in 2012, and continued to undergo checkups (aged 40-74 years). We examined the risk factors for new-onset CKD and 1.5-fold increase in serum creatinine levels among laboratory values from 2012 to 2020. RESULTS: Age, body mass index (BMI), triglyceride levels, atrial fibrillation, and medication for hypertension (HT) and diabetes mellitus were independent risk factors for proteinuria, whereas current smoking, BMI, systolic blood pressure (SBP), and medication for HT were independent risk factors for estimated glomerular filtration rate < 60 mL/min/1.73 m2. SBP, triglyceride levels and medication for HT were risk factors for a 1.5-fold increase in serum creatinine levels during course of the study. The cut-off values of BMI for eGFR < 60 mL/min/1.73 m2 were 22.2 (men 24.7, women 22.1) kg/m2 and fasting triglyceride levels for a 1.5-fold increase in serum creatinine level were 171 (men 247, women 170) mg/dL, respectively. CONCLUSIONS: Health checkups provide information to prevent new-onset CKD and worsening of renal function. It is necessary to increase the rate of health checkups and visits to medical institutions after health checkups as well as to use these results for health guidance.

Dynamic diversity of SARS-CoV-2 genetic mutations in a lung transplantation patient with persistent COVID-19.

Numerous SARS-CoV-2 variant strains with altered characteristics have emerged since the onset of the COVID-19 pandemic. Remdesivir (RDV), a ribonucleotide analogue inhibitor of viral RNA polymerase, has become a valuable therapeutic agent. However, immunosuppressed hosts may respond inadequately to RDV and develop chronic persistent infections. A patient with respiratory failure caused by interstitial pneumonia, who had undergone transplantation of the left lung, developed COVID-19 caused by Omicron BA.5 strain with persistent chronic viral shedding, showing viral fusogenicity. Genome-wide sequencing analyses revealed the occurrence of several viral mutations after RDV treatment, followed by dynamic changes in the viral populations. The C799F mutation in nsp12 was found to play a pivotal role in conferring RDV resistance, preventing RDV-triphosphate from entering the active site of RNA-dependent RNA polymerase. The occurrence of diverse mutations is a characteristic of SARS-CoV-2, which mutates frequently. Herein, we describe the clinical case of an immunosuppressed host in whom inadequate treatment resulted in highly diverse SARS-CoV-2 mutations that threatened the patient's health due to the development of drug-resistant variants.

Photodynamic therapy with paclitaxel-encapsulated indocyanine green-modified liposomes for breast cancer.

Background: Photodynamic therapy (PDT) involves the administration of a photosensitizing agent and irradiation of light at an excitation wavelength that damages tumor cells without causing significant damage to normal tissue. We developed indocyanine green (ICG)-modified liposomes in which paclitaxel (PTX) was encapsulated (ICG-Lipo-PTX). ICG-Lipo-PTX accumulates specifically in tumors due to the characteristics of the liposomes. The thermal and photodynamic effects of ICG and the local release of PTX by irradiation are expected to induce not only antitumor effects but also cancer immunity. In this study, we investigated the antitumor effects of ICG-Lipo-PTX in breast cancer. Methods: The antitumor effects of ICG-Lipo-PTX were examined in xenograft model mice subcutaneously implanted with KPL-1 human breast cancer cells. ICG-Lipo-PTX, ICG-Lipo, or saline was administered intraperitoneally, and the fluorescence intensity was measured with a fluorescence imaging system (IVIS). Intratumor temperature, tumor volume, and necrotic area of tumor tissue were also compared. Next, we investigated the induction of cancer immunity in an allogeneic transplantation model in which BALB-MC mouse breast cancer cells were transplanted subcutaneously in the bilateral inguinal region. ICG-Lipo-PTX was administered intraperitoneally, and PDT was performed on only one side. The fluorescence intensity measured by IVIS and the bilateral tumor volumes were compared. Cytokine secretory capacity was also evaluated by ELISPOT assay using splenocytes. Results: In the xenograft model, the fluorescence intensity and temperature during PDT were significantly higher with ICG-Lipo-PTX and ICG-Lipo in tumor areas than in nontumor areas. The fluorescence intensity in the tumor area was reduced to the same level as that in the nonirradiated area after two times of irradiation. Tumor growth was significantly reduced and the percentage of necrotic area in the tumor was higher after PDT in the ICG-Lipo-PTX group than in the other groups. In the allograft model, tumor growth on day 14 in the ICG-Lipo-PTX group was significantly suppressed not only on the PDT side but also on the non-PDT side. In addition, the secretion of interferon-γ and interleukin-2 was enhanced, whereas that of interleukin-10 was suppressed, in the ICG-Lipo-PTX group. Conclusion: The PDT therapy with ICG-Lipo-PTX may be an effective treatment for breast cancer.

Effects of tea, catechins and catechin derivatives on Omicron subvariants of SARS-CoV-2.

The Omicron subvariants of SARS-CoV-2 have multiple mutations in the S-proteins and show high transmissibility. We previously reported that tea catechin (-)-epigallocatechin gallate (EGCG) and its derivatives including theaflavin-3,3'-di-O-digallate (TFDG) strongly inactivated the conventional SARS-CoV-2 by binding to the receptor binding domain (RBD) of the S-protein. Here we show that Omicron subvariants were effectively inactivated by green tea, Matcha, and black tea. EGCG and TFDG strongly suppressed infectivity of BA.1 and XE subvariants, while effect on BA.2.75 was weaker. Neutralization assay showed that EGCG and TFDG inhibited interaction between BA.1 RBD and ACE2. In silico analyses suggested that N460K, G446S and F490S mutations in RBDs crucially influenced the binding of EGCG/TFDG to the RBDs. Healthy volunteers consumed a candy containing green tea or black tea, and saliva collected from them immediately after the candy consumption significantly decreased BA.1 virus infectivity in vitro. These results indicate specific amino acid substitutions in RBDs that crucially influence the binding of EGCG/TFDG to the RBDs and different susceptibility of each Omicron subvariant to EGCG/TFDG. The study may suggest molecular basis for potential usefulness of these compounds in suppression of mutant viruses that could emerge in the future and cause next pandemic.

Factors Influencing Sarcopenic Changes in YUBI-WAKKA Finger-Ring Test Results After One Year: A Retrospective Observational Study.

Background: The YUBI-WAKKA (finger-ring) test was developed and validated as a predictor of sarcopenia, physical disability, and even mortality. We focused on the sarcopenic status and subsequent changes after 1 year using this test and analyzed factors related to these changes. We also examined the robustness of this test by administering it alongside annual checkups held in local clinics. Methods: We conducted the study to investigate the data of the annual checkup of National Health Insurance at primary care clinics in 2017 and 2018 in Tama City. We studied the participants in a retrospective observational way. The participants of the YUBI-WAKKA test were a total of 5,405 and 4,391 residents between the ages of 65 and 74 years. We first compared anthropological biomarkers of two groups, those that had calves larger than their own finger-ring and those that had calves smaller than their own finger-ring. Then, we compared these results to those from the previous year's 1,048 pairs of data. We were particularly interested in changes from larger to smaller, which we defined here as a sarcopenic change, and performed multivariate logistic regression analysis with forced entry methods to determine the related factors. We also calculated the concordant rate of the test results after 1 year. Results: In total, 14% (men) and 16% (women) each year received a positive (sarcopenic) YUBI-WAKKA test result. The factors related to this sarcopenic change in 1 year were low uric acid in men and a low or decreasing body mass index and increased hemoglobin and alanine aminotransferase in women. The concordant rates of the larger, just fit and smaller groups following finger-ring testing were 72.8%, 63.5%, and 52.0%, respectively. Conclusions: We propose that low uric acid and low or reducing body weight were related to low muscle mass and that ALT and Hb may be indirectly related to sarcopenia. These could be predictive factors to determine sarcopenia-prone individuals. The YUBI-WAKKA (finger-ring) test results and their change were considered to be a sustainable and reasonable way to use in the community with small effort to find those who are at high risk for sarcopenia among the elderly population and who may require intervention.

Characterization of SpsQ from Staphylococcus pseudintermedius as an affinity chromatography ligand for canine therapeutic antibodies.

Coagulase-positive Staphylococci express protein A, which binds to host antibodies, to evade the immune system. Taking advantage of its specific binding to antibodies, protein A from Staphylococcus aureus, which is called SpA, is commonly used as an affinity chromatography ligand for human therapeutic antibodies. However, among four canine IgG subclasses (A, B, C, and D), only IgG-B binds to SpA strongly and establishing an efficient and robust purification scheme for canine therapeutic antibodies whose IgG subclass is A, C, or D remains difficult and depends on finding a suitable substitute to SpA. S. pseudintermedius, a major coagulase-positive Staphylococci found in dogs, expresses spsQ gene which is orthologous to S. aureus spa. We hypothesized that to serve S. pseudintermedius to better adapt to the dog immune system, SpsQ would bind to canine IgGs stronger than SpA, making it a better affinity chromatography ligand for canine therapeutic antibodies. To characterize SpsQ, we first determined the spsQ nucleotide sequence from S. pseudintermedius isolates. Based on the identified sequence, we prepared recombinant proteins containing the immunoglobulin-binding domains of SpA (r-SpA) and SpsQ (r-SpsQ) and determined their binding capacity for each canine IgG subclass. The binding capacity of r-SpsQ for IgG-B was almost as high as that of r-SpA. Interestingly, while both r-SpsQ and r-SpA showed no binding to IgG-C, the binding capacity of r-SpsQ for IgG-A and IgG-D was significantly higher than that of r-SpA. Finally, we performed affinity chromatography using r-SpsQ- or r-SpA-immobilized resin and revealed that the recovery rates of IgG-A and IgG-D using r-SpsQ were significantly higher than those using r-SpA. Our findings indicate that SpsQ has a strong potential to be used as an affinity chromatography ligand for canine therapeutic antibodies of subclass A, B, and D.

Sarcopenia and coexistent risk factors detected using the 'Yubi-wakka' (finger-ring) test in adults aged over 65 years in the public annual health check-up in Tama City, Tokyo: a cross-sectional study.

OBJECTIVES: To examine the positive rate of sarcopenia using the 'Yubi-wakka' (finger-ring) test and associated risk factors among adults aged 65 years and older. DESIGN: Cross-sectional study. SETTING: We used the Yubi-wakka test, which has been developed and validated as a predictor of sarcopenia, frailty, disability and mortality. A positive test result is indicated by a smaller calf circumference than the finger-ring. The test was administered during annual health check-ups among residents of Tama City, Japan. PARTICIPANTS: During the 2019 fiscal year, 12 894 individuals aged 65 years and older underwent the Yubi-wakka test at primary care clinics. INTERVENTIONS: Examinees conducted the test themselves in a seated position. They formed a ring around their calf using both thumbs and index fingers and judged whether their calf was larger, the same or smaller than their finger-ring. PRIMARY AND SECONDARY OUTCOME MEASURES: We compared anthropometric and serological data between the positive (smaller calf) and negative (larger calf) test result groups. RESULTS: The positive rate was 15.4% among men and 18.5% among women. The prevalence of a positive result was higher in those aged ≥80 years than in younger age groups in both sexes (men: 22.8%; women: 28.8%). Multivariate logistic regression analysis showed that a diagnosis of metabolic syndrome was a risk factor for detecting a positive test result in women aged 65-74 years (OR 3.445; 95% CI 1.44 to 8.29) and ≥75 years (OR 3.37; 95% CI 1.97 to 5.78). CONCLUSIONS: Because the Japanese population is healthy and lives long, interventions against sarcopenia are important, especially for older adults aged >75 years. The presence of metabolic syndrome may be a risk factor for sarcopenia (as detected by the Yubi-wakka test) and future frailty, and requires closer attention, especially among women.

Biological properties of Staphylococcus virus ΦSA012 for phage therapy.

Staphylococcus virus ΦSA012 has a wide host range and efficient lytic activity. Here, we assessed the biological stability of ΦSA012 against temperature, freeze-thawing, and pH to clinically apply the phage. In addition, inoculation of ΦSA012 through i.p. and i.v. injections into mice revealed that phages were reached the limit of detection in serum and accumulated notably spleens without inflammation at 48 h post-inoculation. Furthermore, inoculation of ΦSA012 through s.c. injections in mice significantly induced IgG, which possesses neutralizing activity against ΦSA012 and other Staphylococcus viruses, ΦSA039 and ΦMR003, but not Pseudomonas viruses ΦS12-3 and ΦR18 or Escherichia viruses T1, T4, and T7 in vitro. Immunoelectron microscopic analysis showed that purified anti-phage IgG recognizes the long-tail fiber of staphylococcus viruses. Although S. aureus inoculation resulted in a 25% survival rate in a mouse i.p. model, ΦSA012 inoculation (i.p.) improved the survival rate to 75%; however, the survival rate of ΦSA012-immunized mice decreased to less than non-immunized mice with phage i.v. injection at a MOI of 100. These results indicated that ΦSA012 possesses promise for use against staphylococcal infections but we should carefully address the appropriate dose and periods of phage administration. Our findings facilitate understandings of staphylococcus viruses for phage therapy.

Indocyanine green conjugated phototheranostic nanoparticle for photodiagnosis and photodynamic therapy.

BACKGROUND: Phototheranostics represents a highly promising paradigm for cancer therapy, although selecting an appropriate optical imager and sensitizer for clinical use remains challenging. METHODS: Liposomally formulated phospholipid-conjugated indocyanine green, denoted as LP-iDOPE, was developed as phototheranostic nanoparticle and its cancer imaging-mediated photodynamic reaction, defined as the immune response induced by photodynamic and photothermal effects, was evaluated with a near-infrared (NIR)-light emitting diode (LED) light irradiator. RESULTS: Using in vivo NIR fluorescence imaging, we demonstrated that LP-iDOPE was selectively delivered to tumor sites with high accumulation and a long half-life. Following low-intensity NIR-LED light irradiation on the tumor region of LP-iDOPE accumulated, effector CD8+ T cells were activated at the secondary lymphoid organs, migrated, and subsequently released cytokines including interferon-γ and tumor necrosis factor-α, resulting in effective tumor regression. CONCLUSIONS: Our anti-cancer strategy based on tumor-specific LP-iDOPE accumulation and low-intensity NIR-LED light irradiation to the tumor regions, i.e., photodynamic reaction, represents a promising approach to noninvasive cancer therapy.

First isolation of ST398 methicillin-resistant Staphylococcus aureus carrying staphylococcal cassette chromosome mec type IVd from pig ears in Japan.

The emergence and increasing prevalence of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) are a global concern. To investigate the prevalence and characteristics of sequence type 398 (ST398) MRSA in pig ears, 102 pig's ears were collected from 102 animals shipped from 51 farms at an abattoir. Eight ST398 MRSA isolates were isolated from the ears of eight pigs shipped from seven farms. Of the eight ST398 isolates, seven had the staphylococcal cassette chromosome mec (SCCmec) type IVd and these were obtained from seven pigs shipped from six farms. Single nucleotide polymorphisms ranging from 13 to 26 were observed in the core-genome regions in the seven SCCmec type IVd isolates. We believe that this is the first report on the isolation of ST398 MRSA SCCmec type IVd in Japan.

Current status and future perspective of antimicrobial-resistant bacteria and resistance genes in animal-breeding environments.

The emergence and spread of antimicrobial-resistant bacteria (ARB) and antimicrobial resistance genes (ARGs) are a global public health concern. ARB are transmitted directly or indirectly from animals to humans. The importance of environmental transmission of ARB and ARGs has recently been demonstrated, given the relationships between compost, livestock wastewater, insects, and wildlife. In addition, companion animals and their surrounding environments (veterinary hospitals and homes with companion animals) should be considered owing to their close relationship with humans. This review discusses the current status and future perspectives of ARB and ARGs in animal-breeding environments.

Inhibitory effects and gene expression analysis of chemotherapeutic photodynamic therapy by using a liposomally formulated indocyanine green derivative.

BACKGROUND: Photodynamic therapy (PDT) utilizes the enhanced permeability retention effect of photosensitizers and is less invasive and more selective than traditional chemotherapy. We constructed a chemotherapeutic PDT (chemo-PDT) nanoscale drug delivery system using a liposomally formulated indocyanine green derivative (ICG-Lipo) that encapsulated carboplatin and docetaxel (ICG-Lipo-C&D). METHODS: The antitumor effect of chemo-PDT mediated by ICG-Lipo-C&D was evaluated in a murine colon 26 CDF1 mouse model. Gene expression in tumor tissues was analyzed by RNA sequencing. RESULTS: Chemo-PDT using ICG-Lipo-C&D demonstrated an even stronger PDT-enhancing effect than did ICG-Lipo due to the synergistic effect of carboplatin and docetaxel. In addition, gene expression analysis showed that PDT with ICG-Lipo-C&D increased the expression of immune-related genes and decreased the expression of cytoskeleton-related genes. CONCLUSIONS: Chemo-PDT using ICG-Lipo as a photosensitizer as well as a drug delivery system with an enhanced permeability retention effect may be a promising cancer therapy.

Decreasing the abundance of tetracycline-resistant Escherichia coli in pig feces during nursery using flavophospholipol as a pig feed additive.

Tetracyclines (TCs) are widely used for livestock, and the high prevalence of TC-resistant Escherichia coli in livestock has become a serious concern worldwide. In Japan, the National Action Plan on Antimicrobial Resistance in 2016 aimed to reduce the TC resistance rate in E. coli derived from livestock. Flavophospholipol (FPL), used as a feed additive, has an inhibitory effect on the spread of plasmid-mediated antimicrobial resistance. The number of TC-resistant E. coli was determined in pigs administered TCs and/or FPL to clarify the effect of FPL on reducing the number of TC-resistant E. coli in pigs. TC-resistant E. coli and their plasmids were then analyzed. The pigs were divided into four groups: control, doxycycline (DOXY; a TC), FPL, and a DOXY-FPL combination. Their feces were collected from the nursing period to the day before being transported to the slaughterhouse, followed by estimation of TC-resistant E. coli (colony-forming units [CFU]/g). The number of TC-resistant E. coli increased with the use of DOXY, suggesting that DOXY administration provides a selective pressure for TC-resistant E. coli. Supplementation with FPL as a feed additive significantly suppressed the increase in the number of TC-resistant E. coli, especially during the DOXY administration period. Transfer and growth inhibition analyses were performed for TC-resistant isolates. FPL inhibited the conjugational transfer and growth of a few TC-resistant E. coli isolates. These results suggest that FPL is effective against the spread of TC-resistant E. coli.

Anti-microbial resistance of Salmonella isolates from raw meat-based dog food in Japan.

BACKGROUND: Salmonella contamination of raw meat-based diets (RMBDs) for pets poses a major public health concern but has not been investigated in Japan. OBJECTIVE: To investigate Salmonella contamination in RMBDs for dogs marketed in Japan and the anti-microbial resistance profiles of the Salmonella isolates. METHODS: Sixty commercial RMBD samples were collected in the Okayama and Osaka Prefectures, Japan, between December 2016 and March 2017. The obtained Salmonella isolates were serotyped, their anti-microbial resistance patterns were determined, and the anti-microbial-resistant isolates were screened for the presence of resistance genes by polymerase chain reaction. RESULTS: Salmonella enterica subsp. enterica was detected in seven of the 60 RMBD samples. Among them, five isolates were identified as S. Infantis (n = 3), S. Typhimurium (n = 1) and S. Schwarzengrund (n = 1), while the serotypes of two isolates were unable to be identified. All isolates were susceptible to ampicillin, cefazolin, cefotaxime and gentamycin. Two isolates were resistant to more than one anti-microbial agent; one of the S. Infantis isolates was resistant to streptomycin, kanamycin, tetracycline and trimethoprim, while the S. Typhimurium isolate was resistant to nalidixic acid, ciprofloxacin and chloramphenicol. The S. Schwarzengrund isolate was resistant to tetracycline. Additionally, the S. Typhimurium isolate harboured the anti-microbial resistance gene gyrA with a mutation corresponding to Ser-83→Phe amino acid substitution. CONCLUSION: The study findings suggest that RMBDs for dogs marketed in Japan can be a potential source of Salmonella infection for dogs and humans including infections caused by quinolone-resistant isolates.

Broad-host-range IncW plasmid harbouring tet(X) in Escherichia coli isolated from pigs in Japan.

OBJECTIVES: Tetracyclines are used in veterinary medicine for livestock. Tigecycline, a semisynthetic tetracycline derivative, is a last-resort antimicrobial used to treat multidrug-resistant Gram-negative bacterial infections. The prevalence of variants of the mobile tigecycline resistance gene tet(X) in livestock is increasing worldwide. However, the prevalence of Tet(X) among livestock in Japan is unclear. This study was conducted to clarify the prevalence of Tet(X) in pigs in Japan, focusing on isolation and molecular characterisation of plasmid-mediated tet(X)-positive Escherichia coli through retrospective analysis. METHODS: We retrospectively screened for tigecycline-resistant E. coli strains isolated from pigs. The tigecycline-resistant strain and tet(X)-harbouring plasmid were characterised. RESULTS: The IncW plasmid harbouring the tet(X) variant [previously named as tet(X6)] was detected in one E. coli isolate from pigs (0.8%; 1/120) in 2012. The tet(X) plasmid was transferable by conjugation to the E. coli ML4909 recipient strain. Some mobile genetic elements (TnAs3 and ISVsa3) were observed in the region surrounding tet(X). The tet(X)-harbouring plasmid shared a conserved backbone with IncW plasmid R388, which is a broad-host-range plasmid. CONCLUSION: The emergence and spread of tet(X) variants in Enterobacterales poses a public-health concern. To the best of our knowledge, this is the first report of the emergence of an IncW plasmid harbouring tet(X). Using tetracyclines in livestock exerts selective pressure on the tet(X) plasmid; therefore, prudent use of tetracyclines is required.

Campylobacter Express Resistance Array for detecting the presence of fluoroquinolone- and macrolide-resistant Campylobacter jejuni and Campylobacter coli in broiler farms.

AIMS: The aim of the study was to develop a microarray-based method for the detection of antibiotic-resistant Campylobacter in broiler farms to decrease the risk of contamination of chicken meat. METHODS AND RESULTS: A combination of DNA microarray and primer extension for rapid and simultaneous detection of fluoroquinolone- and macrolide-resistant Campylobacter jejuni/Campylobacter coli, termed Campylobacter Express Resistance Array (CAMERA), was used to analyse chicken caecal droppings. CAMERA assays could detect at least 105 colony forming units of C. jejuni/C. coli g-1 of chicken caecal contents spiked with C. jejuni/C. coli. To compare the CAMERA method and direct culturing method for screening antibiotic-resistant C. jejuni/C. coli in poultry farms, chicken caecal droppings obtained from 42 poultry houses were analysed using both methods. In total, 95.2% of the results (40/42 poultry houses) obtained using the CAMERA and culturing method were identical. In the remaining two poultry houses, the CAMERA could detect the prevalent strain of C. jejuni/C. coli based on results of the culturing method. CONCLUSIONS: The culturing method required >3 days to isolate and identify antibiotic-resistant C. jejuni/C. coli. In contrast, the CAMERA required only 6 h. SIGNIFICANCE AND IMPACT OF THE STUDY: This method can facilitate quick screening and control of fluoroquinolone- and macrolide-resistant C. jejuni/C. coli in broiler farms.

Isolation of Human Lineage, Fluoroquinolone-Resistant and Extended-ß-Lactamase-Producing Escherichia coli Isolates from Companion Animals in Japan.

An increase in human and veterinary fluoroquinolone-resistant Escherichia coli is a global concern. In this study, we isolated fluoroquinolone-resistant E. coli isolates from companion animals and characterized them using molecular epidemiological analysis, multiplex polymerase chain reaction to detect E. coli ST131 and CTX-M type extended-spectrum ß-lactamases (ESBL), and multi-locus sequence typing analysis. Using plain-CHROMagar ECC, 101 E. coli isolates were isolated from 34 rectal swabs of dogs and cats. The prevalence of resistance to fluoroquinolone and cefotaxime was 27.7% and 24.8%, respectively. The prevalence of fluoroquinolone-resistant isolates (89.3%) was higher when CHROMagar ECC with CHROMagar ESBL supplement was used for E. coli isolation. The prevalence of cefotaxime resistance was also higher (76.1%) when 1 mg/L of ciprofloxacin-containing CHROMagar ECC was used for isolation. The cefotaxime-resistant isolates possessed CTX-M type ß-lactamase genes (CTX-M-14, CTX-M-15, or CTX-M-27). Seventy-five percent of fluoroquinolone-resistant isolates were sequence types ST131, ST10, ST1193, ST38, or ST648, which are associated with extensive spread in human clinical settings. In addition, we isolated three common fluoroquinolone-resistant E. coli lineages (ST131 clade C1-M-27, C1-nM27 and ST2380) from dogs and their respective owners. These observations suggest that companion animals can harbor fluoroquinolone-resistant and/or ESBL-producing E. coli, in their rectums, and that transmission of these isolates to their owners can occur.

Characterization of Reproductive Microbiota of Primiparous Cows During Early Postpartum Periods in the Presence and Absence of Endometritis.

Endometritis has a major impact on fertility in postpartum dairy cows. Since previous studies showed an association between reproductive microbiota and perinatal disease, we monitored both bovine uterine and vaginal microbiota in primiparous cows to elucidate the effect of early postpartum microbiota on endometritis. Uterine and vaginal samples were collected at time points from pre-calving to 35 days postpartum (DPP), and analyzed by 16S rRNA sequencing, combined with ancillary bacterial culture. A total of seven healthy cows and seven cows diagnosed with endometritis on 35 DPP were used in the current study. The uterine and vaginal microbiota showed a maximum of 20.1% shared amplicon sequence variants (ASVs) at linked time points. 16S rRNA based analysis and traditional culture methods revealed that Trueperella showed a higher abundance in both uterus and vagina of the endometritis group compared to the healthy group on 21 DPP (U-test p < 0.05). Differential abundance analysis of the uterine microbiota showed that Enterococcus and six bacterial genera including Bifidobacterium were unique to the healthy group on the day of calving (0 DPP) and 28 DPP, respectively. In contrast, Histophilus and Mogibacteriaceae were characteristic bacteria in the vagina pre-calving in cows that later developed endometritis, suggesting that these bacteria could be valuable to predict clinical outcomes. Comparing the abundances of bacterial genera in the uterine microbiota, a negative correlation was observed between Trueperella and several bacteria including Lactobacillus. These results suggest that building an environment where there is an increase in bacteria that are generally recognized as beneficial, such as Lactobacillus, may be one possible solution to reduce the abundance of Trueperella and control endometritis.

Current Status of Antimicrobial Drug Use in Japanese Companion Animal Clinics and the Factors Associated With Their Use.

The Japanese National Action Plan on Antimicrobial Resistance (AMR) was adopted to strengthen AMR surveillance and monitoring in companion animals. The Japanese Veterinary Antimicrobial Resistance Monitoring (JVARM) system monitors the sale of veterinary antimicrobial drugs by pharmaceutical companies, and the sale of human drugs by principal wholesale companies to companion animal (dogs and cats) clinics. However, the data do not include sales by local drug suppliers and personal importation to companion animal clinics in Japan. The purposes of this study were to estimate total antimicrobial drug use by companion animal clinics in Japan and to identify the factors associated with their use. In 2018, questionnaires gathering data on attributes of the clinic and volumes of antimicrobial drugs used were sent to 212 clinics across Japan by the Japan Veterinary Medical Association. Out of the clinics, 170 valid questionnaires were returned (80.2% response rate). Antimicrobial drugs were categorized first as human, veterinary, or imported drugs and then further categorized as important drugs (critically important drugs for humans and second-choice veterinary drugs) or others. Total antimicrobial drug use was estimated based on the number of clinics reported in 2016. The relationships between antimicrobial drug use and various questionnaire items were analyzed using non-parametric regression analysis. Total antimicrobial drug use was estimated at 29.9t, which was 2.1 times higher than reported by the JVARM survey on the sales of antimicrobial drugs. In terms of total use, important drugs and human drugs accounted for 12.6 and 61.8%, respectively. Clinic income per veterinarian was associated with total antimicrobial use per veterinarian. The proportion of important drugs among all antimicrobial drugs used in a clinic was high in recently established clinics with middle-aged and older directors.

Aerobic Composting and Anaerobic Digestion Decrease the Copy Numbers of Antibiotic-Resistant Genes and the Levels of Lactose-Degrading Enterobacteriaceae in Dairy Farms in Hokkaido, Japan.

Efficient methods for decreasing the spread of antimicrobial resistance genes (ARGs) and transfer of antimicrobial-resistant bacteria (ARB) from livestock manure to humans are urgently needed. Aerobic composting (AC) or anaerobic digestion (AD) are widely used for manure treatment in Japanese dairy farms. To clarify the effects of AC and AD on antimicrobial resistance, the abundances of antimicrobial (tetracycline and cefazolin)-resistant lactose-degrading Enterobacteriaceae as indicator bacteria, copy numbers of ARGs (tetracycline resistance genes and ß-lactamase coding genes), and concentrations of residual antimicrobials in dairy cow manure were determined before and after treatment. The concentration of tetracycline/cefazolin-resistant lactose-degrading Enterobacteriaceae was decreased over 1,000-fold by both AC and AD. ARGs such as tetA, tetB, and bla TEM were frequently detected and their copy numbers were significantly reduced by ∼1,000-fold by AD but not by AC. However, several ARG copies remained even after AD treatment. Although concentrations of the majority of residual antimicrobials were decreased by both AC and AD, oxytetracycline level was not decreased after treatment in most cases. In addition, 16S rRNA gene amplicon-based metagenomic analysis revealed that both treatments changed the bacterial community structure. These results suggest that both AC and AD could suppress the transmission of ARB, and AD could reduce ARG copy numbers in dairy cow manure.