Suspected Cat-to-Human Transmission of SARS-CoV-2, Thailand, July-September 2021.

A veterinarian in Thailand was diagnosed with COVID-19 after being sneezed on by an infected cat owned by an infected patient. Genetic study supported the hypothesis of SARS-CoV-2 transmission from the owner to the cat, and then from the cat to the veterinarian.

Nationwide Surveillance and Molecular Characterization of Critically Drug-Resistant Gram-Negative Bacteria: Results of the Research University Network Thailand Study.

A large-scale surveillance is an important measure to monitor the regional spread of antimicrobial resistance. We prospectively studied the prevalence and molecular characteristics of clinically important Gram-negative bacilli, including Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii complex (ABC), and Pseudomonas aeruginosa, from blood, respiratory tract, urine, and sterile sites at 47 hospitals across Thailand. Among 187,619 isolates, 93,810 isolates (50.0%) were critically drug resistant, of which 12,915 isolates (13.8%) were randomly selected for molecular characterization. E. coli was most commonly isolated from all specimens, except the respiratory tract, in which ABC was predominant. Prevalence of extended-spectrum cephalosporin resistance (ESCR) was higher in E. coli (42.5%) than K. pneumoniae (32.0%), but carbapenem-resistant (CR)-K. pneumoniae (17.2%) was 4.5-fold higher than CR-E. coli (3.8%). The majority of ESCR/CR-E. coli and K. pneumoniae isolates carried blaCTX-M (64.6% to 82.1%). blaNDM and blaOXA-48-like were the most prevalent carbapenemase genes in CR-E. coli/CR-K. pneumoniae (74.9%/52.9% and 22.4%/54.1%, respectively). In addition, 12.9%/23.0% of CR-E. coli/CR-K. pneumoniae cocarried blaNDM and blaOXA-48-like. Among ABC isolates, 41.9% were extensively drug resistant (XDR) and 35.7% were multidrug resistant (MDR), while P. aeruginosa showed XDR/MDR at 6.3%/16.5%. A. baumannii was the most common species among ABC isolates. The major carbapenemase gene in MDR-A. baumannii/XDR-A. baumannii was blaOXA-23-like (85.8%/93.0%), which had much higher rates than other ABC species. blaIMP, blaVIM, blaOXA-40-like, and blaOXA-58-like were also detected in ABC at lower rates. The most common carbapenemase gene in MDR/XDR-P. aeruginosa was blaIMP (29.0%/30.6%), followed by blaVIM (9.5%/25.3%). The findings reiterate an alarming situation of drug resistance that requires serious control measures.

In vitro synergistic activity of fosfomycin in combination with other antimicrobial agents against carbapenem-resistant Klebsiella pneumoniae isolated from patients in a hospital in Thailand.

INTRODUCTION: Carbapenem-resistant Klebsiella pneumoniae (CRKP) causes high morbidity and mortality worldwide. The purpose of the study was to assess the synergistic activity of fosfomycin in combination with other antimicrobial agents against CRKP isolated from patients in Songklanagarind Hospital, Thailand. METHODS: A total of 35 K. pneumoniae isolates were obtained from patients in Songklanagarind Hospital. The MICs of imipenem and meropenem were determined in all isolates by broth microdilution. In all CRKP isolates, the presence of carbapenemase and extended-spectrum ß-lactamase (ESBL) genes was investigated by PCR, while the production of these enzymes was determined by combined disk test. In the carbapenemase-genes-negative CRKP isolates, the porin loss and efflux pump were characterized by SDS-PAGE and broth microdilution, respectively. Finally, the synergistic effects of fosfomycin and other antimicrobial agents were evaluated by checkerboard analysis. RESULTS: Twenty-one of 35 K. pneumoniae isolates were classified as CRKP. Most of CRKP isolates carried blaNDM-1 (n = 18), blaSHV (n = 21), blaCTX-M (n = 21), and blaTEM (n = 16). In fosfomycin-based combination, the result showed that the highest synergistic activity in this study was observed in the combination of fosfomycin and gentamicin (61.9%). CONCLUSION: These findings suggested that the fosfomycin and gentamicin combination might be useful as a possible treatment option for CRKP infection.

Potential of antimicrobial topical gel with synthesized biogenic silver nanoparticle using Rhodomyrtus tomentosa leaf extract and silk sericin.

OBJECTIVE: Silver nanoparticles synthesized using Rhodomyrtus tomentosa leaf extract and silk sericin were used to functionalize carbopol 940 gel for topical applications. RESULTS: UV-vis spectra presented surface plasmon resonance at 426 nm, transmission electron microscopy revealed that nanoparticles were spherical with an average size of 25-50 nm. X-ray diffraction presented crystalline silver nanoparticles with zeta potential of ≈ - 30 mV. FTIR spectra showed a reduction of silver nitrate indicated by the shift in -OH at 2958 cm-1. The silver nanoparticle demonstrated broad-spectrum antimicrobial activity against Gram-positive, Gram-negative and fungi with MIC ranging between 0.26 and 2.10 µg mL-1, respectively. MIC of hydrogel ranged between 1.05-2.10 µg mL-1 with cell viability of 89%. Spreadability and extrudability of gel were 9.3 ± 0.85 s and 19.85 ± 0.03%, respectively with first order of fickian diffusion. CONCLUSIONS: The silver nanoparticle gel exhibited an effective antimicrobial property, hence can be exploited for topical applications.

Adjunctive therapy of intravenous colistin to intravenous tigecycline for adult patients with non-bacteremic post-surgical intra-abdominal infection due to carbapenem-resistant Acinetobacter baumannii.

Post-surgical intra-abdominal infections (IAIs) due to carbapenem-resistant Acinetobacter baumannii (CRAB) are difficult to treat due to suboptimal peritoneal penetrations of several antimicrobial agents. Tigecycline has favorable outcomes of treating IAIs due to multidrug-resistant organisms but occurrence of breakthrough bacteremia has been observed because this agent has low serum level. Colistin has in vitro activity against CRAB but data on treatment of IAIs is limited due to poor peritoneal penetration. The purpose of this retrospective study is to explore the outcomes of adjunctive intravenous (IV) colistin to IV tigecycline in the treatment of IAIs caused by CRAB. Of 28 patients with non-bacteremic post-surgical IAIs due to CRAB, 14 patients received IV tigecycline alone and 14 patients received IV tigecycline with IV colistin. The 14-day, 30-day, in-hospital mortality rates, the rate of breakthrough bacteremia and the rate of bacterial eradication were not significantly different. The adjunctive therapy of IV colistin was associated with significantly higher rates of renal complications (10/14) than those receiving IV tigecycline alone (3/14) (P value = 0.023). In addition, the patients receiving adjunctive IV colistin had significantly more unfavorable non-clinical outcomes including longer length of hospital stay (P value = 0.049) and higher antimicrobial cost (P value = 0.008) and non-antimicrobial costs (P value = 0.037). In this study, adjunctive IV colistin to conventional IV tigecycline in the treatment of non-bacteremic post-surgical IAIs caused by CRAB did not yield clinical benefit but caused higher renal complication and unfavorable non-clinical outcomes.

Diagnostic scoring system for tuberculous meningitis among adult patients with non-suppurative and non-bacterial meningitis.

Tuberculous meningitis (TBM) is the most severe form of extra-pulmonary tuberculosis. The definite diagnosis of this disease is difficult and can result in delayed treatment. Conventional culture yields low sensitivity while high-sensitivity diagnostic techniques are costly and unpractical. Adenosine deaminase (ADA) is used to diagnose several settings of extra-pulmonary tuberculosis but it is limited in TBM especially among HIV-infected patients. We retrospectively reviewed the data of patients with non-suppurative meningitis and compared the patient data with TBM and other causes including carcinomatous, lymphomatous, lymphocytic and fungal meningitis. We found that HIV infection, diabetes mellitus, duration of symptoms <14 days, radiologic findings of hydrocephalus, and CSF ADA level >10 IU were associated with TBM. The scoring system based on these parameters and their coefficients in the final model achieved an area under the receiver operating characteristic curve of 0.95,625. The indices were HIV infection = 5, diabetes mellitus = 3, duration of symptoms <14 days = 5, hydrocephalus = 4, and ADA in CSF >10 IU = 5. Based on the assumed costs of the patients with false negative and false positive, an appropriate cut off value of 10 was selected and the sensitivity was 92% and specificity was 89%.

Characterization of a Chikungunya virus strain isolated from banked patients' sera.

BACKGROUND: Chikungunya virus (CHIKV) is a prevalent mosquito-borne pathogen that is emerging in many parts of the globe causing significant human morbidity. Here, we report the isolation and characterization of an infectious CHIKV from banked serum specimens of suspected patients from the 2009 epidemic in Thailand. METHODS: Standard plaque assay was used for CHIKV isolation from the banked serum specimens. Isolated CHIKV was identified base on E1 structural gene sequence. Growth kinetic, infectivity, cell viability and cytokine gene expression throughout CHIKV infection in a permissive cell line, 293T cells, was performed using several approaches, including standard plaque assay, immunofluorescence assay, classical MTT assay, and quantitative real-time PCR. Two tailed Student's t test was used for evaluation statistically significance between the mean values of the groups. RESULTS: Based on the E1 structural gene sequence and phylogenetic analysis, we identified the virus as the CHIK/SBY8/10 isolate from Indonesia. Assessment of the growth kinetics, cytopathic effects as well as its ability to induce cellular immune responses suggested that the currently isolated CHIK/SBY8/10 virus is relatively more virulent than a known CHIKV vaccine strain, which also induces more dramatic proinflammatory responses. CONCLUSIONS: Our studies further add to the infectivity of a less-studied yet infectious CHIKV isolate as well as underscored the importance and utility of 293T cells as an excellent cell culture model for studying viral growth, CHIKV-induced inflammatory cellular responses and cell death. Together, these studies provide novel information on the CHIKV biology, infectivity and virus-cell interaction, which would help develop novel interventions against the infection.

Early detection of novel Leishmania species DNA in the saliva of two HIV-infected patients.

BACKGROUND: Leishmaniasis caused by two new species of Leishmania; L. siamensis and L. martiniquensis have been recently described in Thailand. The disease has mainly been documented in AIDS patients from southern Thailand. In this study, polymerase chain reaction (PCR) was used to determine HIV-Leishmania co-infection in southern Thailand. METHODS: One ml of saliva and 3 ml of EDTA blood were collected from HIV-infected patients for PCR detection of Leishmania DNA, cloning and sequencing. The positive PCR samples were then cultured on Schneider's insect medium. RESULTS: Three out of 316 saliva samples collected from HIV-infected patients were found to be positive for Leishmania DNA (0.95%). Among the positive samples, one patient was observed with disseminated cutaneous lesions and also tested positive via saliva, whole blood and buffy coat in PCR. The second case presenting with nodular lesions also gave a positive saliva test via PCR two months prior to buffy coat. This diagnosis was confirmed by microscopic examination and a culture of biopsy samples from a nodule. The last case was an asymptomatic Leishmania infection which tested PCR positive only in saliva with a consecutive sample collection conducted for three months. CONCLUSIONS: The prevalence of Leishmania infection in HIV infected patients within this study is 0.95%. Leishmania DNA was detected in saliva by PCR prior to blood and buffy coat of two HIV infected patients. Early detection of Leishmania DNA in saliva would be beneficial for the follow up of asymptomatic Leishmania infected patients, the early treatment of leishmaniasis and for surveillance survey purpose. However, full evaluation of sensitivity and specificity of this technique with a large cohort of patients is required before deployment.

Clinical Outcomes of Community-Acquired Severe Sepsis after Implementation of a Simple Severe Sepsis Fast Track.

Background: Severe sepsis and septic shock are the most common causes of in-hospital death in Songkhla Provincial Hospital and half of the patients are transferred from community hospitals. A simple severe sepsis and septic shock fast track had been implemented in all community hospitals in Songkhla Province and in Songkhla Provincial Hospital in December 2013. Objective: Evaluate the clinical outcomes and predictors of mortality of severe sepsis and septic shock patients after implementation of the simple severe sepsis and septic shock fast track. Material and Method: A retrospective study of all available medical records between December 2013 and May 2014 of hospitalized patients aged older than 15 years with a final diagnosis of severe sepsis or septic shock. Results: Of 723 patients diagnosed as community acquired sepsis, 228 (31.5%) patients were diagnosed with severe sepsis or septic shock. A simple severe sepsis and septic shock fast track were activated in 69.3%. Patients in the activated fast track group had significantly lower mortality than the non-activated fast track group (21.0% vs. 42.9%, p = 0.001). After adjusted analysis, the four independent risk factors associated with increased mortality in severe sepsis and septic shock were initial presentation with systolic blood pressure lower than 90 mmHg (adjusted odds ratio [aOR] 2.57), central nervous system failure (aOR 7.33), acute renal failure (aOR 5.07), and received norepinephrine (aOR 2.87). Two factors associated with a significant decrease in mortality were the simple fast track activated at the emergency department (aOR 0.22) or at the ward (aOR 0.09) and received appropriate initial antibiotics (aOR 0.09). Conclusion: Early recognition and early resuscitation in case of severe sepsis and septic shock can reduce mortality. A simple severe sepsis and septic shock fast track should be implemented in all community hospitals. It is a simple clinical diagnosis with simple management that is possible in every community hospital before transfer to a secondary or tertiary care hospital.

Detection of chikungunya virus antigen by a novel rapid immunochromatographic test.

Chikungunya fever is a mosquito-borne disease of key public health importance in tropical and subtropical countries. Although severe joint pain is the most distinguishing feature of chikungunya fever, diagnosis remains difficult because the symptoms of chikungunya fever are shared by many pathogens, including dengue fever. The present study aimed to develop a new immunochromatographic diagnosis test for the detection of chikungunya virus antigen in serum. Mice were immunized with isolates from patients with Thai chikungunya fever, East/Central/South African genotype, to produce mouse monoclonal antibodies against chikungunya virus. Using these monoclonal antibodies, a new diagnostic test was developed and evaluated for the detection of chikungunya virus. The newly developed diagnostic test reacted with not only the East/Central/South African genotype but also with the Asian and West African genotypes of chikungunya virus. Testing of sera from patients suspected to have chikungunya fever in Thailand (n = 50), Laos (n = 54), Indonesia (n = 2), and Senegal (n = 6) revealed sensitivity, specificity, and real-time PCR (RT-PCR) agreement values of 89.4%, 94.4%, and 91.1%, respectively. In our study using serial samples, a new diagnostic test showed high agreement with the RT-PCR within the first 5 days after onset. A rapid diagnostic test was developed using mouse monoclonal antibodies that react with chikungunya virus envelope proteins. The diagnostic accuracy of our test is clinically acceptable for chikungunya fever in the acute phase.

Impact of antibiotic exposure on occurrence of nosocomial carbapenem-resistant Acinetobacter baumannii infection: a case control study.

Carbapenem-resistant Acinetobacter baumannii (CRAB) infection is one of the most important healthcare associated diseases worldwide. Although antibiotic use is recognized as a risk factor for CRAB infection, the impact of antibiotic class and length of use on CRAB infection is still unclear. A case-control study was conducted in adult intensive care units and general wards of Songklanagarind Hospital, a tertiary-care hospital in southern Thailand, to investigate the effect of different antibiotic exposure and the duration of use on the risk of developing CRAB infection. Cases were defined as patients with carbapenem-susceptible A. baumannii (CSAB) or CRAB infection. Controls were randomly selected from patients and matched 1:1 with cases using ward and date of admission. Multinomial logistic regression was used to compute relative risk ratios (RRR) and 95% confidence intervals (CI) for CRAB infection. Of 197 cases with A. baumannii infection, there were 139 with CRAB infection and 58 with CSAB infection. Compared to the control group, use of fluoroquinolones, broad-spectrum cephalosporins and carbapenems for more than three days increased the risk of CRAB infection with RRR (95% CI) of 81.2 (38.1-862.7), 31.3 (9.9-98.7) and 112.1 (7.1-1770.6), respectively. The RRR (95% CI) for one to three day treatment of fluoroquinolones, broad-spectrum cephalosporins and carbapenems were 5.4 (0.8-38.7), 6.2 (0.1-353.2) and 63.3 (15.6-256.9), respectively. Long-term use of certain antibiotics and even short term use of carbapenems increased the risk of CRAB infection. In this setting, use of these antibiotics, especially carbapenems, should be limited to reduce CRAB infection.

Clinical outcomes of hospital-acquired infection with Acinetobacter nosocomialis and Acinetobacter pittii.

The role of Acinetobacter nosocomialis and Acinetobacter pittii, which belong to the A. calcoaceticus-A. baumannii complex, in hospital-acquired infections is increasingly recognized. Here we describe a retrospective cohort study of hospital-acquired A. calcoaceticus-A. baumannii complex infections at a university hospital in Thailand. A total of 222 unique cases were identified between January 2010 and December 2011. The genomospecies of the A. calcoaceticus-A. baumannii complex isolates were classified as follows: A. baumannii, 197 (89%); A. nosocomialis, 18 (8%); and A. pittii, 7 (3%). All A. nosocomialis and A. pittii isolates were susceptible to imipenem and meropenem. The patients infected with A. nosocomialis and A. pittii had lower 30-day mortality than those infected with carbapenem-susceptible A. baumannii (P = 0.025) and carbapenem-resistant A. baumannii (P = 0.013). The factors influencing 30-day mortality were infection with non-baumannii A. calcoaceticus-A. baumannii complex (hazard ratio [HR], 0.12; 95% confidence interval [CI], 0.03 to 0.51; P = 0.004), infection with carbapenem-resistant A. baumannii (HR, 1.57; 95% CI, 0.89 to 2.79; P = 0.105), appropriate empirical antimicrobial therapy (HR, 0.38; 95% CI, 0.23 to 0.61; P < 0.001), and higher acute physiology and chronic health evaluation II (APACHE II) score (HR, 1.15; 95% CI, 1.10 to 1.19; P < 0.001). In Galleria mellonella assays, the survival rates were significantly higher for the larvae infected with A. nosocomialis or A. pittii than for those infected with either carbapenem-susceptible A. baumannii or carbapenem-resistant A. baumannii, but no differences in survival rates were observed between carbapenem-susceptible A. baumannii and carbapenem-resistant A. baumannii. These findings suggest intrinsic differences in virulence between non-baumannii A. calcoaceticus-A. baumannii complex species and A. baumannii but not between carbapenem-susceptible and resistant A. baumannii.

Single dosage of doxycycline for prophylaxis against leptospiral infection and leptospirosis during urban flooding in southern Thailand: a non-randomized controlled trial.

This study was conducted to investigate the protective efficacy of a single dosage of 200 mg doxycycline against leptospiral infection and leptospirosis and associated risk factors among residents exposed to flooding in southern Thailand. Of 641 participants, 600 received doxycycline while 41 did not. Twenty two participants were infected with Leptospira and six developed leptospirosis. Having a laceration wound was significantly associated with leptospiral infection (odds ratio [OR] = 37.20; P < 0.001) and leptospirosis (OR = 18.24; P = 0.003) whereas exposure to flood more than 3 h per day was associated with only leptospiral infection (OR = 3.70; P = 0.038). Seventeen participants who received doxycycline and five who did not, were infected with Leptospira, resulting a protective efficacy of 76.8% (95% confidence interval [CI] = 34.3%-92.0%). Four who received doxycycline and two who did not, developed leptospirosis, resulting a protective efficacy of 86.3% (CI = -9.8%-98.2%). Among the participants with laceration wound, the protective efficacy for leptospiral infection was 92.0% (CI = 81.2%-96.6%) and for leptospirosis was 95.6% (CI = 78.2%-99.3%). Among the participants exposed to flood water less than or equal to 3 h per day, the protective efficacy for leptospiral infection was 89.2% (95% CI 63.6%-96.67%). A single dosage of 200 mg doxycycline for prophylaxis might be effective for preventing leptospirosis among flood victims with laceration wound after recent flood exposure.

Animal reservoirs and potential vectors of Leishmania siamensis in southern Thailand.

Leishmania siamensis is newly described as the causative pathogen of autochthonous leishmaniasis in Thailand. Potential vectors and animal reservoirs of L. siamensis are not thoroughly studied. An environmental survey was carried out in the affected area in two provinces in southern Thailand: Songkhla and Nakhon Si Thammarat. Ninety-nine villagers, 378 sandflies, and potential animal reservoirs were examined. Leishmania DNA amplicon was identified in two species of female sandflies, Sergentomyia (Neophlebotomus) and Sergentomyia (Parrotomyia) barraudi. The DNA amplicon was also identified in black rats (Rattus rattus). A phylogenetic tree of confirmed patients, sandflies and black rats fell into a single clade and separate from other Leishmania species. This study showed the potential involvement of R. rattus and Sergentomyia (Neophlebotomus and Parrotomyia) sandflies in transmission of L. siamensis.

Antibacterial and Anti-Biofilm Efficacy of Endolysin LysAB1245 against a Panel of Important Pathogens.

Infections caused by antibiotic-resistant bacteria pose a significant global challenge. This study explores the antibacterial effects of a bacteriophage-derived endolysin, LysAB1245, against important pathogens, including Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Staphylococcus aureus. We determined the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) for all tested isolates. A time-kill study was conducted to evaluate the reduction in bacterial survival following treatment with LysAB1245. Additionally, the effects of LysAB1245 on P. aeruginosa K1455 and methicillin-resistant S. aureus (MRSA) NPRC 001R-formed biofilms were investigated. The MIC and MBC of LysAB1245 against all the tested isolates ranged from 4.68 to 9.36 µg/mL and 4.68 to 18.72 µg/mL, respectively. The time-kill study demonstrated more than a 4 log CFU/mL (99.99%) reduction in bacterial survival within 6 h of LysAB1245 treatment at 2MIC. LysAB1245 (1/8-1/2MIC) treatment significantly reduced biofilms formed by P. aeruginosa and MRSA in a concentration-dependent manner. Furthermore, scanning electron and confocal laser scanning microscopy confirmed the potential inhibition effects on 3-day established biofilms formed on abiotic surfaces upon treatment with LysAB1245 at 2MIC. The findings indicate that endolysin LysAB1245 could be employed as a new alternative therapeutic antibacterial and anti-biofilm agent for combating biofilm-related infections.

Comparative immunogenicity and neutralizing antibody responses post heterologous vaccination with CoronaVac (Sinovac) and Vaxzevria (AstraZeneca) in HIV-infected patients with varying CD4+ T lymphocyte counts.

The immune response to heterologous coronavirus disease (COVID-19) vaccination in people living with HIV (PLWH) is still unclear. Herein, our prospective cohort study aimed to compare the immune response of heterologous vaccination with CoronaVac (Sinovac) and Vaxzevria (AstraZeneca) between PLWH having CD4 counts ≤ 200 cells/µL (low CD4+) and > 200 cells/µL (high CD4+). Anti-receptor-binding domain (RBD) immunoglobulin G (IgG) levels and the percentage inhibition of neutralizing antibodies (nAbs) were analyzed at 2 and 12 weeks after immunization. Participants in the low and high CD4+ groups had mean CD4+ counts of 139 and 575 cell/µL, respectively. Two and 12 weeks after immunization, in the low CD4 group, the median anti-RBD-IgG levels were 159 IU/mL and 143 IU/mL, respectively, whereas the nAb level was 71% and decreased to 47.2%, respectively. Contrarily, the median anti-RBD-IgG levels in the high CD4+ group were 273 IU/mL and 294 IU/mL, respectively, whereas the nAb levels were 89.3% and relatively stable at 81.6%. However, although immune responses between the two study groups were not significantly different, a decline in nAb levels was observed at 12 weeks in the low CD4+ group. Therefore, a COVID-19 booster vaccine dose is suggested for immunoprotection.

Genotypic and phenotypic mechanisms underlying antimicrobial resistance and synergistic efficacy of rifampicin-based combinations against carbapenem-resistant Acinetobacter baumannii.

Purpose: Carbapenem-resistant Acinetobacter baumannii (CRAB) is an urgent concern to public health. This study focuses on exploring the resistance mechanisms and the in vitro results of using rifampicin in combination with conventional antibiotics for the management of CRAB. Methods: The synergistic and bactericidal effects of rifampicin with conventional antibiotics were evaluated using chequerboard assay and time-kill assay, while the phenotypic and genotypic characteristics of resistant determinants were performed by efflux pump detection and whole genome sequencing on 29 isolates from ICU patients with underlying health diseases. Results: The isolates showed multidrug resistance, with over 60% showing addictive responses to rifampicin-based combinations at FICI ranging from 0.6 to 0.8. The time-kill assay revealed 99 % killing for rifampicin and minocycline combination in one isolate at 1/4 MIC rifampicin plus 1/4 MIC minocycline, while a bacteriostatic effect was observed at 1/2 MIC rifampici plus 1/2 MIC for a second isolate. Combination with tigecycline resulted in a 99% killing in two out of three isolates with a 2.5-3 log reduction in CFU at 1/4 MIC rifampicin plus 1/4 MIC tigecycline. Rifampicin plus colistin exhibited bactericidal activity against three out of four isolates. The combinations of rifampicin with ciprofloxacin, chloramphenicol, and trimethoprim-sulfamethoxazole were ineffective against the isolates. In addition, a 4-fold reduction in rifampicin MIC was observed in 2 out of 14 isolates in the presence of an efflux pump inhibitor. The pan-genome study demonstrated a progressive evolution with an accessory genome estimated to cover 58% of the matrix. Seven of the ten sequenced isolates belong to sequence type 2 (ST2), while one isolate each was assigned to ST164, ST16, and ST25. Furthermore, 11 plasmids, 34 antimicrobial resistance (AMR) genes, and 65 virulence-associated genes were predicted from the whole genome data. The blaOXA-23blaADC-25, blaOXA-66, blaPER-7, aph(6)-Id, armA, and arr-3 were prevalent among the isolates. Sequence alignment of the bacteria genome to the reference strain revealed a deleterious mutation in the rpoB gene of 4 isolates. Conclusion: The study suggests that rifampicin in combination with either minocycline, tigecycline, or colistin might be a treatment option for CRAB clinical isolates. In addition, genotypic analysis of the bacteria isolates may inform the clinician of the suitable drug regimen for the management of specific bacteria variants.

Genomic Characterization of Multidrug-Resistant Enterobacteriaceae Clinical Isolates from Southern Thailand Hospitals: Unraveling Antimicrobial Resistance and Virulence Mechanisms.

The emergence and spread of antimicrobial resistance (AMR) among Enterobacteriaceae pose significant threats to global public health. In this study, we conducted a short-term surveillance effort in Southern Thailand hospitals to characterize the genomic diversity, AMR profiles, and virulence factors of Enterobacteriaceae strains. We identified 241 carbapenem-resistant Enterobacteriaceae, of which 12 were selected for whole-genome sequencing (WGS) and genome analysis. The strains included Proteus mirabilis, Serratia nevei, Klebsiella variicola, Klebsiella aerogenes, Klebsiella indica, Klebsiella grimontii, Phytobacter ursingii, Phytobacter palmae, Kosakonia spp., and Citrobacter freundii. The strains exhibited high levels of multidrug resistance, including resistance to carbapenem antibiotics. Whole-genome sequencing revealed a diverse array of antimicrobial resistance genes (ARGs), with strains carrying genes for ß-lactamase, efflux pumps, and resistance to other antibiotic classes. Additionally, stress response, metal tolerance, and virulence-associated genes were identified, highlighting the adaptability and pathogenic potential of these strains. A plasmid analysis identified several plasmid replicons, including IncA/C2, IncFIB(K), and Col440I, as well as several plasmids identical to those found globally, indicating the potential for the horizontal gene transfer of ARGs. Importantly, this study also identified a novel species of Kosakonia spp. PSU27, adding to the understanding of the genetic diversity and resistance mechanisms of Enterobacteriaceae in Southern Thailand. The results reported in this study highlight the critical importance of implementing effective antimicrobial management programs and developing innovative treatment approaches to urgently tackle AMR.

Association of biofilm formation, antimicrobial resistance, clinical characteristics, and clinical outcomes among Acinetobacter baumannii isolates from patients with ventilator-associated pneumonia.

INTRODUCTION: Biofilm formation is an important virulence factor of Acinetobacter baumannii. Here, we examined the biofilm formation of archived A. baumannii causing ventilator-associated pneumonia (VAP). METHODS: Eighteen and twenty isolates of A. baumannii causing bacteremic pneumonia and non-bacteremic pneumonia were included, respectively. Antimicrobial susceptibility testing was performed by broth microdilution method, while biofilm formation was evaluated by microtiter dish biofilm formation assay. RESULTS: All 38 isolates were still susceptible to colistin and tigecycline, whereas almost all isolates were non-susceptible (intermediate to resistant) to several antimicrobial agents, especially ceftriaxone and cefotaxime. Approximately, 44% of bacteremic isolates and 50% of non-bacteremic isolates were classified as carbapenem-resistant A. baumannii (CRAB). Biofilm formation was detected in 42% of the studied isolates. Bacteremia among the patients infected with biofilm-producing isolates was significantly higher than in those infected with non-biofilm-producing isolates. The antimicrobial susceptibilities of A. baumannii with biofilm formation were lower than those without biofilm formation, but the differences did not have statistical significance. The patients infected with non-biofilm-producing isolates had good clinical and non-clinical outcomes than those infected with biofilm-producing isolates. The survival rate of patients diagnosed with VAP due to biofilm-producing A. baumannii was lower than in those patients diagnosed with VAP due to non-biofilm-producing isolates. CONCLUSION: Biofilm formation of A. baumannii causing VAP was associated with antimicrobial resistance and bacteremia as well as unfavorable clinical outcomes.

Characterization of a novel bacteriophage endolysin (LysAB1245) with extended lytic activity against distinct capsular types associated with Acinetobacter baumannii resistance.

Capsular polysaccharides are considered as major virulence factors associated with the ability of multidrug-resistant (MDR) Acinetobacter baumannii to cause severe infections. In this study, LysAB1245, a novel bacteriophage-encoded endolysin consisting of a lysozyme-like domain from phage T1245 was successfully expressed, purified, and evaluated for its antibacterial activity against distinct capsular types associated with A. baumannii resistance. The results revealed a broad spectrum activity of LysAB1245 against all clinical MDR A. baumannii isolates belonging to capsular type (KL) 2, 3, 6, 10, 47, 49, and 52 and A. baumannii ATCC 19606. At 2 h following the treatment with 1.7 unit/reaction of LysAB1245, more than 3 log reduction in the numbers of bacterial survival was observed. In addition, LysAB1245 displayed rapid bactericidal activity within 30 min (nearly 3 log CFU/mL of bacterial reduction). Thermostability assay indicated that LysAB1245 was stable over a broad range of temperature from 4 to 70°C, while pH sensitivity assay demonstrated a wide range of pH from 4.5 to 10.5. Furthermore, both minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of LysAB1245 against all MDR A. baumannii isolates and A. baumannii ATCC 19606 were 4.21 µg/mL (0.1 unit/reaction). Conclusively, these results suggest that LysAB1245 possesses potential application for the treatment of nosocomial MDR A. baumannii infections.