Multidrug Resistance Related to Biofilm Formation in Acinetobacter baumannii and Klebsiella pneumoniae Clinical Strains from Different Pulsotypes.

The emergence of Acinetobacter baumannii and Klebsiella pneumoniae strains in the hospital environment has been associated with the presence of multiple genetic elements, virulence factors and the ability to form biofilms. This study evaluated the biofilm formation ability of clinical and environmental A. baumannii and K. pneumoniae strains, isolated from various sources and presenting different molecular characteristics, resistance profiles and pulsed-field gel electrophoresis patterns. Fifty-three isolates were recovered from 2009 to 2014 in a Brazilian university hospital. Investigation of biofilm formation was performed for 10 strains of each species assessed by an initial adhesion assay, biofilm cell concentration and biofilm biomass, evaluated by quantitative assays in replicates, in three independent experiments. All strains of A. baumannii were able to attach to polystyrene plates, although two strains adhered to a lesser degree than the control. K. pneumoniae strains showed opposite behaviour, where only three strains adhered significantly when compared to the control. Quantitative evaluation revealed that in five A. baumannii and four K. pneumoniae isolates the biomass production could be characterised as moderate. None of the isolates were strong biofilm producers. Our results demonstrate: (1) biofilm formation is a heterogeneous property amongst A. baumannii and K. pneumoniae clinical strains and it was not associated with certain clonal types; (2) no relationship between multidrug resistance and biofilm production was observed; (3) more virulent K. pneumoniae strains tended to present higher production of biofilm.

A sustained endemic outbreak of vancomycin-resistant Enterococcus faecium: A 30-month surveillance study.

BACKGROUND: The assessment of risk factors for the nosocomial acquisition of colonization and infection by vancomycin-resistant Enterococcus faecium (VREfm) is often problematic due to scarce data on antibiotic use. A 30-month prospective cohort study was conducted to characterize VREfm strains isolated during an outbreak and endemic period, identifying the risk factors, antibiotic consumption, and prevalence of virulence determinants. METHODS: The study was conducted in a tertiary care hospital. A representative number (171 patients) of isolates that were classified as resistant to high-level vancomycin (minimum inhibitory concentration (MIC) ≥ 256 µg/ml) were investigated. RESULTS: Among 171 colonized patients, 22 (12.9%) developed VRE infection. All VREfm isolates harboured vanA genes. Genes codifying virulence factors such as enterococcal surface protein (esp), aggregation substance 1 (asa1), and gelatinase (gelE) were detected in the VREfm studied. All patients infected with VRE had previously been colonized and became infected on average 14 days after colonization. Only previous use of aminoglycosides was a risk factor independently associated with VRE infection; however, glycopeptide consumption in defined daily doses (DDD) per 1000 patient-days was associated with the presence of this microorganism. The monthly colonization pressure ranged from 0.004% to 1.32% during the 30-month study period. CONCLUSIONS: We found a high incidence of VRE in a tertiary care hospital, independently associated with the prior use of aminoglycosides and the administration of glycopeptides.

Relationship between antimicrobial use and the highest number of multidrug-resistant-Pseudomonas aeruginosa: a 10-year study.

INTRODUCTION: Multi-drug-resistant (MDR) Pseudomonas aeruginosa is a dangerous pathogen causing nosocomial infection, particularly in low- and middle-income countries like Brazil. This retrospective study at a Brazilian university hospital examined the relationship between antimicrobial use and MDR-P. aeruginosa. METHODOLOGY: Data was collected from 358 patients with non-repetitive P. aeruginosa infections from 2009 to 2019. Antibiotic use was measured in grams and expressed as defined daily dose (DDD) per 1000 patient-days for meropenem, imipenem, polymyxin, and tigecycline. RESULTS: Extensively drug-resistant (XDR) P. aeruginosa occurred in 36.1%, and MDR in 32.6% of cases. Risk factors for XDR infection were hospitalization prior to infection (OR = 0.9901), intensive care unit (ICU) admission (OR = 0.4766), previous antibiotic use (OR = 1.4417), and use of cefepime (OR = 0.3883). Over the ten-year period, utilization of the monitored antibiotics increased, and there was a positive correlation between the rise in MDR-P. aeruginosa and the consumption of ceftriaxone, imipenem, meropenem, and polymyxin B. The 30-day mortality rate was 40.0% for all patients and 41.0% for those infected with XDR-P. aeruginosa. CONCLUSIONS: This study highlights the negative impact of the indiscriminate use of antimicrobials, which has led to a significant increase in multidrug-resistant P. aeruginosa strains in hospitals.

Effectiveness of indonesian honey on the acceleration of cutaneous wound healing: an experimental study in mice.

The purpose of this study was to investigate the effectiveness of Indonesian honey in wound healing compared to Tegaderm hydrocolloid dressing and Manuka honey. Three groups of male mice were treated to produce 2 circular, full-thickness skin wounds on the dorsum. They were then randomly allocated to receive daily Indonesian honey, Manuka honey, or hydrocolloid (control). Macroscopic findings were observed from day 0 to 14 after wounding. Microscopic findings on days 3, 7, 11, and 14 after wounding were obtained. The ratios of wound areas for honey groups on day 3 were smaller than those of the control group. Wound areas of honey groups gradually decreased to almost the same wound area as the control group on day 14, while the control group wound area peaked on day 5 and rapidly decreased until day 14. On day 3, myofibroblasts and new blood capillaries in wound tissue of honey groups were observed, but were not seen in the control group. After day 7, microscopic findings were almost the same among the 3 groups. These results indicate that Indonesian honey is almost as effective for wound healing as Manuka honey and hydrocolloid dressing. .

Pressure ulcers colonized by MRSA as a reservoir and risk for MRSA bacteremia in patients at a brazilian university hospital.

UNLABELLED: Pressure ulcers (PU) are a major reservoir of methicillin-resistant Staphylococcus aureus (MRSA) in hospitals. The objectives of this study were to estimate the prevalence of MRSA colonization in PU of hospitalized patients with Stage II or higher PU, to identify risk factors for colonization of these wounds, and to ascertain whether MRSA colonization of PU increases the risk of MRSA bacteremia. METHODS: This study was conducted at the Clinical Hospital of Federal University of Uberlandia, MG, Brazil. A prospective cohort study of 145 patients with Stage II or higher PU, colonized or not with MRSA, was conducted over 21 months. Infected ulcers were defined for clinical signs and for positive evaluation of smears of the wound by the ratio of polymorphonuclears to epithelial cells of ≥ 2:1, after Giemsa staining. RESULTS: Sixty-three (43.5%) MRSA colonized PU patients were identified, but none of the risk factors analyzed were independently associated with MRSA colonization. Among the patients with positive blood cultures and MRSA colonized PU, the odds ratio for MRSA bacteremia (OR = 19.0, 95% CI = 2.4-151.1, P < 0.001) and mortality rate (OR = 21.9, 95% CI = 1.23-391.5, P = 0.002), were high. Independent risk factors for MRSA bacteremia were: ≥ 2 underlying disease (OR = 6.26, 95% CI = 1.01-39.1, P = 0.05) and prior MRSA infected PU (OR = 12.75, 95% CI=1.22-132.9, P = 0.03). CONCLUSION: The present study identifies MRSA colonized PU patients as a potential epidemiologic reservoir for this organism and a high-risk for MRSA bacteremia, which contributes to prolonged hospitalization and poor prognosis. .

Genetic Alterations Associated with Polymyxin B Resistance in Nosocomial KPC-2-Producing Klebsiella pneumoniae from Brazil.

The rapid increased multidrug resistance in Klebsiella pneumoniae has led to a renewed interest in polymyxin antibiotics, such as colistin, as antibiotics of last resort, not least in low/middle income countries. We conducted a genomic survey of clinical polymyxin-resistant K. pneumoniae to investigate the genetic alterations in isolates harboring blaKPC-2. Whole-genome sequencing was performed using an Illumina NextSeq 500 paired-end reads. Mutations and insertion sequence detection were analyzed to seven isolates recovered from clinical specimens of patients hospitalized in Brazil, focusing on key genes associated with polymyxin resistance. Furthermore, the levels of mRNA expression of genes associated with resistance to polymyxin B and other antimicrobials were evaluated by quantitative real-time PCR. Eighty-five percent of the isolates were assigned to clonal complex 258, with a minimum inhibitory concentration range of 4 to >256 mg/L for polymyxin B. It was possible to observe the presence of one important insertion element, ISKpn13, in a strain recovered from the blood that have blaKPC-2. Deleterious mutations reported in PmrB (R256G), YciM (N212T), and AcrB (T598A) were common, and mobile colistin resistance (mcr) genes were absent in all the isolates. RT-qPCR analysis revealed an overexpression of the pmrC (1.160-fold), pmrD (2.258-fold), and kpnE (1.530-fold) genes in the polymyxin B-resistant isolates compared with the expression of the polymyxin B-susceptible K. pneumoniae isolate. Overall, these results demonstrate the diversity of genetic variations in polymyxin-resistant populations derived from the different clonal strains, but the same sequence types, and suggest that there are still unknown mechanisms of polymyxin resistance in K. pneumoniae.

Novel ST1465/CC216 Nosocomial Lineage of Carbapenem-Resistant Acinetobacter baumannii Harboring an Unusual Plasmid Carrying blaNDM-1 Gene.

This study used whole-genome sequencing to analyze the first case of NDM-1-producing Acinetobacter baumannii belonging to the novel sequence type 1465/CC216 recovered in Brazil. The study identified an unusual plasmid carrying blaNDM-1 gene, in which some genes of the Tn125 transposon were lost. Besides, on the chromosome, the strain reported here presented blaOXA-106 gene, a variant of blaOXA-51 gene, and blaADC-25 with ISAba1 upstream. The isolation of new STs of A. baumannii carrying blaNDM-1 genes elicits our concerns about the possible spread of these genes among clinically relevant bacteria.

High mortality by nosocomial infections caused by carbapenem-resistant P. aeruginosa in a referral hospital in Brazil: facing the perfect storm.

Introduction. Carbapenem-resistant Pseudomonas aeruginosa is responsible for increased patient mortality.Gap Statement. Five and 30 day in-hospital all-cause mortality in patients with P. aeruginosa infections were assessed, followed by evaluations concerning potential correlations between the type III secretion system (TTSS) genotype and the production of metallo-ß-lactamase (MBL).Methodology. This assessment comprised a retrospective cohort study including consecutive patients with carbapenem-resistant infections hospitalized in Brazil from January 2009 to June 2019. PCR analyses were performed to determine the presence of TTSS-encoding genes and MBL genes.Results. The 30-day and 5-day mortality rates for 262 patients were 36.6 and 17.9 %, respectively. The unadjusted survival probabilities for up to 5 days were 70.55 % for patients presenting exoU-positive isolates and 86 % for those presenting exo-negative isolates. The use of urinary catheters, as well as the presence of comorbidity conditions, secondary bacteremia related to the respiratory tract, were independently associated with death at 5 and 30 days. The exoS gene was detected in 64.8 % of the isolates, the presence of the exoT and exoY genes varied and exoU genes occurred in 19.3 % of the isolates. The exoU genotype was significantly more frequent among multiresistant strains. MBL genes were not detected in 92 % of the isolates.Conclusions. Inappropriate therapy is a crucial factor regarding the worse prognosis among patients with infections caused by multiresistant P. aeruginosa, especially those who died within 5 days of diagnosis, regardless of the genotype associated with TTSS virulence.

Incidence of infections caused by carbapenem-resistant Acinetobacter baumannii.

BACKGROUND: Carbapenem-resistant Acinetobacter baumannii (CR-Ab) has become a worrying health care problem, mainly in developing countries, such as Brazil. The objective was to investigate the prevalence and prognostic factors for CR-Ab infections at a Brazilian university hospital and examine the impact of inappropriate antimicrobial therapy on patient outcome. METHODS: A retrospective study on hospitalized patients with CR-Ab infections was carried out from January 2013 to December 2017. An epidemiologic analysis was carried out to determine the frequency of infections, the epidemiologic indicators by year, the risk factors for 30-day mortality, and the impact of inappropriate therapy. RESULTS: A total of 489 patients were included in the study. A rate of 0.7 per 1,000 patient-day CR-Ab infections was observed, mostly in the lungs (54.7%), and predominantly in the adult intensive care unit. The occurrence of infections by CR-Ab per 1,000 patient-days in November 2014 exceeded the established control limit, confirming an outbreak. CONCLUSIONS: The prevalence of CR-Ab increased in the investigated hospital, passing to an endemic pathogen with a direct impact on mortality and the control of these strains.

Epidemiologically relevant antimicrobial resistance phenotypes in pathogens isolated from critically ill patients in a Brazilian Universitary Hospital.

Antimicrobial resistance is a threat to public health worldwide and is associated with higher mortality and morbidity. Despite the extensive knowledge about this problem, drug resistance has continued to emerge, especially in intensive care units (ICUs). The objective of this study was to evaluate the frequencies of epidemiologically relevant resistance phenotypes in pathogens isolated from ventilator-associated pneumonia (VAP), bloodstream infections (BSI) and urinary tract infections (UTI) in patients admitted in the adult intensive care unit (AICU) of the Clinical Hospital of Federal University of Uberlândia, during an one year period. Additionally, at the period of the study, the antibiotic consumption in AICU was verified. Coagulase-negative staphylococci and S. aureus were the main agents of BSI (43.9%), with 60.0% of oxacilin-resistance for both microorganisms, Klebsiella-Enterobacter group predominated in UTI (23.4%), with resistance to third generation cephalosporins in 58.0% of the isolates; and, Pseudomonas aeruginosa in VAP (42.0%), with 72.0% of resistance to imipenem. Cephalosporins (49.6%), vancomycin (37.4%) and carbapenems (26.6%) were the most prescribed antibiotics in the unit. The comparison of the results with a publication of the NNIS program evidenced a worse situation in the studied hospital, mainly between Gram-negative, that had surpassed the percentile 90% elaborated by that system. Based on these results a reconsideration on the empirical use of antibiotics and on prevention and control of nosocomial infections practices is recommended.

Association of Colistin-Resistant KPC Clonal Strains with Subsequent Infections and Colonization and Biofilm Production.

Carbapenemase-producing organisms are pandemic and a significant threat to public health. We investigated the clonal relatedness of colistin-resistant Klebsiella pneumoniae strains producing KPC-type carbapenemase (KPC-KP) causing subsequent infections or colonization. Moreover, we aimed to gain insight into the ability of biofilm production in K. pneumoniae strains producing carbapenemase. Twenty-two consecutive KPC-KP and one KPC-negative strain was identified from an adult intensive care unit in Brazil. Seventy-five percent of isolates that harbored the blaKPC gene exhibited genetic relatedness by pulsed-field gel electrophoresis, and none presented the plasmid-mediated mcr-1 and blaNDM genes. This study showed that the majority of repeated KPC infections in adults were caused by a clone that caused the previous infections/colonizations even after a long period of time and illustrates the capacity of multiple clones producing biofilms to coexist in the same patient at the same time, becoming a reservoir of KPC-KP in the hospital environment.

Nosocomial bloodstream infections: organisms, risk factors and resistant phenotypes in the Brazilian University Hospital.

Bacteremia is one of the most frequent and challenging hospital-acquired infection and it is associated with high attributable morbidity and mortality and additional use of healthcare resources. The objective of this work was to determine the frequencies of its occurrence, organisms and resistance phenotypes associated to nosocomial acquired bloodstream infections. A total number of 51 nosocomial bacteremia by Gram-negative and 99 by Gram-positive were evaluated and compared during a 15-month period. The risk factors associated with these bacteremias were analyzed and antibiotic use and surgery were associated with bacteremia by Gram-negative and > 2 invasive devices with Gram-positive. The resistance phenotypes ESBL (extended-spectrum beta-lactamases) (23.5%) and AmpC/others (17.6%) correspond to 41.2 % with predominance of E. agglomerans among AmpC (44.4%) and K. pneumoniae among ESBLs (38.5%). Among S. aureus bacteremia, approximately 40% were associated to MRSA (methicillin-resistant Staphylococcus aureus).

Etiology and pathogenesis of bloodstream infections associated with the use of long-term central vascular catheter (CVC) in patients who undergone gastrointestinal surgery.

CVC is the main factor of risk of bloodstream infections. This study purpose was determining both etiology and pathogenesis of these infections in 80 patients who undergone gastrointestinal surgery and who worn long lasting CVC, in the institution HC-UFU. Cultures were made in nostril, skin of the insertion site, tip and catheter hub, in addition to hemoculture in those suspects of sepsis. The colonization incidence rate of the catheter tip was 12.5/ 1,000 catheter days and the CVC associate infection rate was 3.1/1,000 catheter days. Frequencies of skin, hub and catheter tip colonization were 13.8%, 8.9% and 13.3%, respectively. Coagulase-negative Staphylococci were the most found microorganisms in nostril (74%), skin (45.4%) and hub (75%) and Gram-negative bacilli (50%), followed by S. aureus (25%) the most common ones in catheter tip. Approximately 51% of patients received antibiotics and most of them (53.7%) had therapeutic purpose. The frequency of patients with clinical sepsis was 27.5%. Three cases of bacteremia associated with the use of CVC were detected (3.8%), with S. aureus in two of them and K. pneumoniae in the third one. There was not seen any association of skin and hub colonization with their presence in the catheter tip and in the blood of these patients, but S. aureus was recovered from nostril of those with sepsis by this pathogenic agent. A greater concern is suggested over preventive measures and control of these primary and secondary bloodstream, as well as catheter tip colonization.

Vancomycin-resistant VanA phenotype Enterococcus faecalis: first case in Minas Gerais state and epidemiological considerations.

We report the first case of vancomycin-resistant Enterococcus faecalis with VanA phenotype and vanA genotype isolated from a surgical patient in a general Hospital in Uberlândia, state of Minas Gerais, Brazil.

IncX3 plasmid harboring a non-Tn4401 genetic element (NTEKPC) in a hospital-associated clone of KPC-2-producing Klebsiella pneumoniae ST340/CG258.

IncX-type plasmids have achieved clinical significance for their contribution in the dissemination of genes confering resistance to carbapenems (most blaKPC- and blaNDM-type genes) and polymyxins (mcr-type genes), both antibiotics considered last resort for multidrug-resistant Gram-negative infections. In this study, we report the identification and complete sequence analysis of an IncX3 plasmid (designated pKP1194a) carrying a non-Tn4401 genetic element (NTEKPC) of tnpR-tnpA (partial)-blaKPC-2-ΔISKpn6/traN, originating from a hospital-associated lineage of K. pneumoniae belonging to the ST340/CG258, with epidemiological link to Brazil.

Spread of multidrug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa clones in patients with ventilator-associated pneumonia in an adult intensive care unit at a university hospital.

BACKGROUND: In Brazil, ventilator-associated pneumonia (VAP) caused by carbapenem resistant Acinetobacter baumannii and Pseudomonas aeruginosa isolates are associated with significant mortality, morbidity and costs. Studies on the clonal relatedness of these isolates could lay the foundation for effective infection prevention and control programs. OBJECTIVES: We sought to study the epidemiological and molecular characteristics of A. baumannii vs. P. aeruginosa VAP in an adult intensive care unit (ICU). METHODS: It was conducted a cohort study of patients with VAP caused by carbapenem resistant A. baumannii and P. aeruginosa during 14 months in an adult ICU. Genomic studies were used to investigate the clonal relatedness of carbapenem resistant OXA-23-producing A. baumannii and P. aeruginosa clinical isolates. The risk factors for acquisition of VAP were also evaluated. Clinical isolates were collected for analysis as were samples from the environment and were typed using pulsed field gel electrophoresis. RESULTS: Multivariate logistic regression analysis identified trauma diagnosed at admission and inappropriate antimicrobial therapy as independent variables associated with the development of A. baumannii VAP and hemodialysis as independent variable associated with P. aeruginosa VAP. All carbapenem resistant clinical and environmental isolates of A. baumannii were OXA-23 producers. No MBL-producer P. aeruginosa was detected. Molecular typing revealed a polyclonal pattern; however, clone A (clinical) and H (surface) were the most frequent among isolates of A. baumannii tested, with a greater pattern of resistance than other isolates. In P. aeruginosa the most frequent clone I was multi-sensitive. CONCLUSION: These findings suggest the requirement of constant monitoring of these microorganisms in order to control the spread of these clones in the hospital environment.

Molecular epidemiological survey of the quinolone- and carbapenem-resistant genotype and its association with the type III secretion system in Pseudomonas aeruginosa.

This study evaluated the predictors of mortality and the impact of inappropriate therapy on the outcomes of patients with bacteraemia and ventilator-associated pneumonia (VAP). Additionally, we evaluated the correlation of the type III secretion system (TTSS) effector genotype with resistance to carbapenems and fluoroquinolones, mutations in the quinolone resistance-determining regions (QRDRs), metallo-ß-lactamase and virulence factors. A retrospective cohort was conducted at a tertiary hospital in patients with multidrug-resistant (MDR) P. aeruginosa bacteraemia (157 patients) and VAP (60 patients). The genes for blaIMP, blaVIM, blaSIM, blaGIM and blaSPM and virulence genes (exoT, exoS, exoY, exoU, lasB, algD and toxA) were detected; sequencing was conducted for QRDR genes on fluoroquinolone-resistant strains. The multivariate analyses showed that the predictors independently associated with death in patients with bacteraemia were cancer and inappropriate therapy. Carbapenem resistance was more frequent among strains causing VAP (53.3 %), and in blood we observed the blaSPM genotype (66.6 %) and blaVIM genotype (33.3 %). The exoS gene was found in all isolates, whilst the frequency was low for exoU (9.4 %). Substitution of threonine to isoleucine at position 83 in gyrA was the most frequent mutation among fluoroquinolone-resistant strains. Our study showed a mutation at position 91 in the parC gene (Glu91Lys) associated with a mutation in gyrA (Thre83Ile) in a strain of extensively drug-resistant P. aeruginosa, with the exoT(+)exoS(+)exoU(+) genotype, that has not yet been described in Brazil to the best of our knowledge. This comprehensive analysis of resistance mechanisms to carbapenem and fluoroquinolones and their association with TTSS virulence genes, covering MDR P. aeruginosa in Brazil, is the largest reported to date.

Comparing hospital infections in the elderly versus younger adults: an experience in a Brazilian University Hospital.

The elderly population will grow rapidly over the next 25 years, however there is little information about hospital infections in this group of patients in Brazil. We examined the prevalence of nosocomial and community infections in elderly ((3) 65 years) patients and their relationship with intrinsic and extrinsic risk factors in a Brazilian University Hospital. A total of 155 hospitalized elderly patients were evaluated, and clinical and demographic information about each patient was obtained from hospital records. The rates of nosocomial and community infections were 16.1% and 25.6%, respectively. When the elderly group with and without nosocomial infections was analyzed, practically all the risk factors considered (use of antibiotics, invasive devices, surgery and time of hospitalization) were significantly more associated with the patients with hospital infection. All patients with nosocomial infections were taking antibiotics and most of them (56.0%) were being treated with two or more antibiotics; the length of hospitalization was double (p=0.007) compared to patients who had not acquired hospital infection. The most frequent sites of nosocomial and community infections were surgical (56.0%) and the skin (37.1%), and most of the patients (47.5%) were in the surgical clinic wards. In conclusion, the elderly patients were more likely to develop a nosocomial infection (16.1% prevalence). Surgical infection accounted for the majority (56.0%) of the nosocomial infections, in contrast with North American studies that indicate urinary tract infections to be the commonest.