Colonization by multidrug-resistant bacteria in hematological patients undergoing hematopoietic stem cell transplantation and clinical outcomes: A single-center retrospective cohort study.

BACKGROUND: Bloodstream infections are a leading cause of death in patients who undergo hematopoietic stem cell transplantation (HSCT) and are more severe when caused by multidrug-resistant (MDR) bacteria. This study proposed to investigate if colonization by MDR bacteria negatively affects the clinical outcomes in hematological patients after HSCT, as well as to evaluate possible risk factors for death due to bacteremia by the same colonizing agent. METHODS: A single-center retrospective cohort study was conducted with 405 hematological patients submitted to a single HSCT procedure between 2015 and 2021. Patients were classified as colonized (n = 132) or noncolonized (n = 273) based on the surveillance cultures from D-30 to D+30 of transplantation, and their relevant clinical and laboratory data were collected until D+100. RESULTS: Colonization by MDR bacteria increased blood culture positivity by all micro-organisms and also specifically by MDR bacteria, with a more pronounced effect when caused by carbapenemase-producing Klebsiella pneumoniae. Patients colonized with carbapenem-resistant K. pneumoniae had increased overall mortality (HR = 4.07, 95% CI 1.85-8.91, P = .0005) and had prolonged hospital length of stay in the context of autologous transplantation. Risk factors for death due to bacteremia by the same colonizing agent were neutropenia, colonization by carbapenem-resistant K. pneumoniae and use of high-dose total body irradiation in conditioning. CONCLUSION: Hematological patients colonized by MDR bacteria presented a higher incidence of bloodstream infections, and colonization by carbapenemase-producing K. pneumoniae was associated with reduced overall survival.

Analysis of Clostridioides difficile Infection in Children with Diarrhea in Two Hospitals in Southern Brazil.

Clostridioides difficile infection (CDI) has been increasingly observed in children, but there is a lack of epidemiological and molecular data on CDI in Latin America. This prospective cohort study aimed to investigate the role of CDI in children with diarrhea. It included 105 children with antimicrobial-associated diarrhea (AAD) and analyzed the molecular characteristics of strains isolated from two hospitals in southern Brazil between 2017 and 2020. Fecal samples from the participants were tested for glutamate dehydrogenase (GDH) and A/B toxins using a rapid enzyme immunoassay. GDH-positive samples underwent automated real-time polymerase chain reaction and toxigenic culture. Toxigenic C. difficile isolates were selected for whole genome sequencing. Out of the 105 patients, 14 (13.3%) met the criteria for CDI. Children with a history of previous CDI and the presence of mucus in their stool were more likely to have CDI. Metronidazole was the most used treatment (71.4%), and three patients (23.1%) experienced CDI recurrence (rCDI). Although the number of sequenced isolates was limited, a wide diversity of sequence types (ST) was observed. In addition to toxin genes (tcdA, tcdB, cdtA, and cdtB), the isolates also exhibited virulence factors involved in adhesion (cwp66, groEL, slpA, fbpA/fbp68) and immune evasion (rmlA, rmlB, rmlC, gnd, rfbA-1), along with multiple resistance factors (gyrA mutation, norA, ermB, dfrF, and vanG). These findings highlight the prevalence and recurrence of CDI among hospitalized children. Longitudinal studies are needed to better understand the characteristics of CDI-associated diarrhea and its impact on the healthcare system in this population.

Prevalence and Seasonality of Clostridiodes difficile over 12 Years at a Tertiary Hospital in Brazil.

Clostridioides difficile is the main pathogen responsible for antibiotic-associated diarrhea in adults. Besides its challenging diagnosis, C. difficile infection (CDI) causes substantial morbidity and mortality. Commercially, there are assays with different targets and performances in sensitivity and specificity. The objectives of this study were to: (1) evaluate the prevalence and seasonal variability of CDI rates at a tertiary hospital in southern Brazil over 12 years and (2) determine the impact of using a two-step algorithm test in the laboratory diagnosis. Between January 2007 and May 2019, fecal samples from 2275 patients were analyzed in a cross-sectional study. Four commercial tests were adopted for the diagnosis of CDI, the immunochromatographic test for toxin A from 2007 to 2010; the enzyme-linked immunosorbent assay method for toxins A and B from 2011 to March 2017; and the rapid enzyme immunoassay (EIA) for GDH and toxins A and B, associated with a Polymerase Chain Reaction (PCR) for the toxin B gene from June 2017 to 2019. The annual prevalence was 8.7% from 2007 to March 2017, increasing between June 2017 and 2019 to 14.7% when the C. diff Quik Chek Complete + GeneXpert C. difficile (two-step algorithm) test was adopted. The number of samples (691) and percentage of CDI cases (10.5%) were higher in winter, but the difference has no statistical significance (P > 0.05). An accurate diagnosis and adequate knowledge of the local seasonality of CDI allow the effective implementation of prevention and control strategies for nosocomial CDI, in addition to effective treatment for patients.

Evaluation of MicroScan WalkAway for Determination of Ceftazidime-Avibactam and Ceftolozane-Tazobactam Susceptibility in Carbapenem-Resistant Gram-Negative Bacilli.

We evaluated the performance of ceftazidime-avibactam and ceftolozane-tazobactam MicroScan Neg multidrug-resistant MIC 1 (NMR1) panel for clinical carbapenem-nonsusceptible Gram-negative bacilli isolates. We evaluated 212 clinically significant carbapenem-nonsusceptible Gram-negative bacilli (139 Pseudomonas aeruginosa and 73 KPC-producing Enterobacterales) from 71 Brazilian hospitals (2013 to 2020). Ceftazidime-avibactam and ceftolozane-tazobactam MICs from the panel were compared with a broth microdilution (BMD) test as the reference method. Essential agreement (EA) and categorical agreement (CA) were assessed. For P. aeruginosa, antimicrobial susceptibility testing error rates were calculated using the error-rate bound method. Discrepancies were initially observed with 11 isolates; 4 resolved after retesting, 2 in favor of the NMR1 and 2 in favor of the BMD method. The ceftazidime-avibactam EA (overall and evaluable) was 100% for P. aeruginosa and Enterobacterales. The CA was 100% for Enterobacterales and 98.6% for P. aeruginosa. The ceftolozane-tazobactam EA was 98.6% and 100% (overall and evaluable, respectively), and the CA was 96.4% for P. aeruginosa. For ceftazidime/avibactam, no very major error (VME) was found, and the major error (ME) rate was 4.2% (2/48). For ceftolozane-tazobactam and P. aeruginosa, using the CLSI breakpoints, the minor error (mE) was 11.4%, and no VME or ME was found. While using EUCAST breakpoints, the VME was 11.4% with no ME. The mE becomes ME or VME in the absence of the intermediate category. All categorical errors were also within 1 log of MIC variation, and the adjusted error rate for CLSI/EUCAST was 0% (0/212). The NMR1 panel is an option to test ceftazidime-avibactam for KPC-producing Enterobacterales and carbapenem-nonsusceptible P. aeruginosa. When a MIC of 4 mg/liter for ceftolozane-tazobactam is obtained using this method, an alert could be created, and the results could be confirmed by an alternative method.

Predictive factors, outcomes, and molecular epidemiology of Clostridioides difficile diarrhea in Brazilian hospitals.

Little is known about the role of lineage of strains of Clostridioides difficile (CD) on the clinical presentation of CD infection (CDI) in Latin America, especially regarding the treatment response. We conducted a multicenter, prospective study to investigate the predictive factors and treatment outcomes of CDI in hospitalized patients and to performed phenotypical and molecular characterization of CD strains. A total of 361 diarrheic patients at 5 hospitals from different regions of the country were enrolled. All stool samples were tested for glutamate dehydrogenase (GDH), toxins A and B, and toxin genes using a nucleic acid amplification test (NAAT). Specimens were cultured and susceptibility profile and whole-genome sequencing (WGS) were performed. CDI positivity was 15% (56/377). Predictive factors for CDI were prior use of meropenem (OR 4.09, 95% CI 2.097-7.095; p<0.001), mucus in stools (OR 3.29; 95% CI 1.406-7.722; p=0.006) and neutrophil left-shift with >20% of bands (OR 3.77; 95% IC 1.280-11.120; p=0.016). Overall mortality was 19%, with no deaths attributed to CDI. Oral metronidazole was used in 74% of cases, with 85% of cure and 14% of recurrence. A total of 35 CD isolates were recovered, all of them susceptible to metronidazole and vancomycin. The WGS revealed 17 different STs, six of which were novel. ST42 was the most common ST and hypervirulent strains were not found. Severe CDI were caused by ST42, ST5, ST8, ST48, ST33 and a novel ST667. The ermB gene was more frequently found in isolates of ST42 (p=0.004).

Multicenter study of the epidemiology of Clostridioides difficile infection and recurrence in southern Brazil.

Clostridioides (Clostridium) difficile is the main etiology underlying antibiotic-associated diarrhea (AAD). Still, few Brazilian data are available on this infection. The aims of this multicenter study were to identify the prevalence, clinical characteristics, and outcomes of C. difficile infection (CDI) in patients with antibiotic associated diarrhea at eight hospitals in Curitiba, southern Brazil, during the years 2017-2019. Stool samples were tested using enzyme immunoassay for glutamate dehydrogenase antigen (GDH) and A/B toxins. Positive GDH samples were further evaluated by real-time polymerase chain reaction (PCR) for the presence of genes encoding toxin B (tcdB), binary toxin (cdt), and marker of hypervirulent C. difficile strain (tcdC deletion). The prevalence of CDI in 351 patients with AAD included in the study was 17.7% (n = 62). Among the CDI cases, tcdB was positive in all 62 stool samples, while cdt was positive in 10 samples, and tcdC deletion was positive in only two. Carriage of carbapenem-resistant Gram-negative bacilli, previous hospitalization, and use of broad-spectrum cephalosporin and carbapenem were associated with CDI. Among patients with CDI, 64.5% presented with severe diarrhea, and 8% (5/62) progressed with colitis and required intensive care. The 30-day mortality was 24% (15/62), and the CDI-associated mortality was 4.8% (3/62). Overall, 83.8% (52/62) of the patients achieved primary cure, and 20.8% of the evaluated patients (10/48) presented CDI recurrence. The treatment administered was not significantly associated with the 60-day recurrence or mortality. In conclusion, we reported in this study data of prevalence and recurrence rates of CDI in patients with AAD and evaluated the number of severe cases and infection-related mortality, which were up to now unknown in Southern Brazilian hospitals.

Prevalence of Clostridioides difficile associated diarrhea in hospitalized patients in five Brazilian centers: A multicenter, prospective study.

Epidemiological data on CD infection (CDI) in Latin American are scarce. CDI prevalence and strains characterization were prospectively evaluated in 5 Brazilian hospitals from different regions. Prevalence rates of CDI were 15%, ranging from 0 to 37%. ST42 was the most common Sequence Type and hypervirulent strains were not identified.

Comparison of Cerebrospinal Fluid Biomarkers for Differential Diagnosis of Acute Bacterial and Viral Meningitis with Atypical Cerebrospinal Fluid Characteristics.

OBJECTIVE: Several cerebrospinal fluid (CSF) biomarkers are used to distinguish between acute bacterial meningitis (BM) and viral meningitis (VM). We compared the ability of lactate and glucose (GL) in CSF and the CSF/blood GL ratio to distinguish between acute BM and VM with typical and atypical CSF characteristics. METHODS: Three hundred and twenty-four CSF reports were included, which were distributed as the acute BM, VM, and normal control groups (n = 63, 139, and 122, respectively). RESULTS: Lactate level in the CSF of acute BM group was 4-fold higher than that in the acute VM and control groups (p < 0.0001). CSF lactate presented higher specificity (92%) and negative predictive value (94%) compared to CSF GL and CSF/blood GL ratio in distinguishing acute BM and VM. Definitive acute BM or VM with atypical CSF cell characteristics was observed in 23.2 and 21.6% of samples, respectively, and these groups showed reduced performance of characteristics of all CSF biomarkers. CSF lactate showed better operational characteristics than those of CSF GL and CSF/blood GL ratio, presenting the highest positive likelihood ratio, and thus aided in the differential diagnosis of VM with atypical CSF. CONCLUSION: The CSF lactate assay can be routinely used in laboratories as a rapid, automated, and easy method that is independent of lactate blood levels.

Validation of Mycobacterium tuberculosis real-time polymerase chain reaction for diagnosis of tuberculous meningitis using cerebrospinal fluid samples: a pilot study.

Background Timely diagnosis of tuberculous meningitis (TBM) remains challenging. Molecular diagnostic tools are necessary, particularly in low- and middle-income countries. There is no approved commercial polymerase chain reaction (PCR) assay that can be used to detect Mycobacterium tuberculosis in non-respiratory samples, such as the cerebrospinal fluid (CSF). We aimed to validate the threshold cycle (Ct) cut-off points; calculate the operational characteristics of real-time PCR for detection of M. tuberculosis (MTb qPCR) in the CSF; and the inhibitory affect of CSF red blood cells (RBC) and total proteins on MTb qPCR. Methods A total of 334 consecutive participants were enrolled. Based on clinical, laboratory and imaging data, cases of suspected TBM were categorized as definite, probable, possible or not TBM cases. Receiver operating characteristic curve analysis was used to select the best discriminating Ct value. Results For TBM cases categorized as definite or probable (n=21), the Ct validated for CSF (≤39.5) improved the diagnostic performance of MTb qPCR on CSF samples. The sensitivity was 29%, specificity was 95%, positive predictive value was 26%, negative predictive value was 95%, efficiency was 90% and positive likelihood was 5.3. The CSF RBC and total protein did not affect the positivity of the MTb qPCR. Conclusions These data support the validation of a highly specific but low sensitive MTb qPCR assay for the TBM diagnosis using CSF samples. MTb qPCR contributes significantly to the diagnosis, mainly when associated with conventional microbiology tests and clinical algorithms.

Characterization of CTX-M enzymes, quinolone resistance determinants, and antimicrobial residues from hospital sewage, wastewater treatment plant, and river water.

Multidrug-resistant (MDR) bacteria are widespread in hospitals and have been increasingly isolated from aquatic environments. The aim of the present study was to characterize extended-spectrum ß-lactamase (ESBL) and quinolone-resistant Enterobacteriaceae from a hospital effluent, sanitary effluent, inflow sewage, aeration tank, and outflow sewage within a wastewater treatment plant (WWTP), as well as river water upstream and downstream (URW and DRW, respectively), of the point where the WWTP treated effluent was discharged. ß-lactamase (bla) genes, plasmid-mediated quinolone resistance (PMQR), and quinolone resistance-determining regions (QRDRs) were assessed by amplification and sequencing in 55 ESBL-positive and/or quinolone-resistant isolates. Ciprofloxacin residue was evaluated by high performance liquid chromatography. ESBL-producing isolates were identified in both raw (n=29) and treated (n=26) water; they included Escherichia coli (32), Klebsiella pneumoniae (22) and Klebsiella oxytoca (1). Resistance to both cephalosporins and quinolone was observed in 34.4% of E. coli and 27.3% of K. pneumoniae. Resistance to carbapenems was found in 5.4% of K. pneumoniae and in K. oxytoca. Results indicate the presence of blaCTX-M (51/55, 92.7%) and blaSHV (8/55, 14.5%) ESBLs, and blaGES (2/55, 3.6%) carbapenemase-encoding resistance determinants. Genes conferring quinolone resistance were detected at all sites, except in the inflow sewage and aeration tanks. Quinolone resistance was primarily attributed to amino acid substitutions in the QRDR of GyrA (47%) or to the presence of PMQR (aac-(6')-Ib-cr, oqxAB, qnrS, and/or qnrB; 52.9%) determinants. Ciprofloxacin residue was absent only from URW. Our results have shown strains carrying ESBL genes, PMQR determinants, and mutations in the gyrA QRDR genes mainly in hospital effluent, URW, and DRW samples. Antimicrobial use, and the inefficient removal of MDR bacteria and antibiotic residue during sewage treatment, may contribute to the emergence and spreading of resistance in the environment, making this a natural reservoir.