Isolation of Klebsiella pneumoniae and Acinetobacter radioresistens from Dish Wash Scrubbers

Aims: To identify predominant microorganisms in dish washing scrubbers collected from ten different sources. Study Design: Collection of dish wash scrubbers from different sources, pour plating of appropriately prepared dish wash scrub suspensions and subsequent isolation and identification of predominant isolates. Assessment of antibiotic susceptibility of the selected isolates by disc diffusion assay. Place and Duration of Study: April, 2018 - June 2018. Methodology: A total of 10 dish wash scrubbers (synthetic green scrubber pads free from any anti-bacterial preservatives belonging to the same brand) were collected from various sources. Appropriately prepared dish wash scrub suspensions in peptone water were pour plated on Plate Count Agar (PCA) and MacConkey agar. Predominant colonies selected from the plates based on the colony morphology were subjected to Grams staining, catalase, oxidase, indole, citrate, urease tests and genotypic identification by 16S ribosomal RNA sequencing. The identified isolates were tested for their susceptibility to eight antibiotics by disc diffusion method. Results: Irrespective of the sample source, most of the dish wash scrubbers sampled harbored similar types of colonies. From the colonies obtained two of them were identified by 16S rRNA sequencing and subsequent blasting as Klebsiella pneumoniae and Acinetobacter radioresistens. The isolates were deposited in the NCBI database with accession numbers MK032217 (Klebsiella pneumoniae RSV02) and MK032134 (Acinetobacter radioresistens RSV 01). These isolates were tested for their susceptibility to different antibiotics and Acinetobacter radioresistens RSV 01 was found to be more antibiotic susceptible than Klebsiella pneumoniae RSV02. Conclusion: Observations of this study confirm the potential role of dish wash scrubbers as vehicle for potential pathogens and their ability to act as cross contaminating agents in food processing environments.

Perfil de susceptibilidade aos carbapenêmicos de bacilos Gram-negativos fermentadores da glicose isolados de hemoculturas / Carbapenem susceptibility profile of Gram-negative glucose fermenting bacilli isolated from hemocultures

As Enterobacteriaceae são bactérias Gram-negativas e frequentes causadoras de infecções hospitalares. Os carbapenêmicos (CROs) são considerados as mais recentes linhas de defesa contra infecções por microrganismos multirresistentes, no entanto o desenvolvimento de bactérias multirresistentes a esta classe medicamentosa tem prejudicado o tratamento farmacológico. Nesse sentido, o presente estudo trata-se de uma análise retrospectiva, tendo como objetivo avaliar o perfil de resistência e susceptibilidade de bacilos Gram-negativos fermentadores da glicose aos carbapenêmicos em isolados de hemoculturas positivas no período de janeiro de 2018 a janeiro 2019. Das 5.733 hemoculturas realizadas nesse período, cerca de 5,4% (311 amostras) foram positivas. Dentre essas, 34,4% (107 amostras) positivas para enterobactérias e 65,6% (204 amostras) positivas para outras espécies. Foi observada uma maior incidência de Klebsiella pneumoniae (34,6% dos casos), seguido pela Klebsiella sp. (28,9%) e a Escherichia coli (26,2%). Contudo, três (03) isolados de hemocultura da espécie Klebsiella pneumoniae se mostraram resistentes aos três antibióticos (ertapenem, meropenem e imipenem). Dois isolados de Enterobacter cloaceae também obtiveram resistência aos antibióticos utilizados e uma espécie de Enterobacter aerogenes mostrou-se resistente apenas ao ertapenem e meropenem e sensibilidade ao imipenem.Enterobacteriaceae are Gram-negative bacteria and frequent causes hospital infections. Carbapenemic (CRO) are considered the latest lines of defense against infections by multidrug-resistant microorganisms, however the development of multidrug-resistant bacteria in this drug class has impaired treatment pharmacological. The present study is a retrospective analysis, with the objective of evaluating the resistance and susceptibility profile from isolates in blood cultures from January 2018 to January 2019, from positive blood cultures with enterobacteria growth and analysis of antibiograms performed from fermenter Gram-negative glucose bacilli to carbapenem antimicrobials. Of the 5,733 blood cultures performed from January 2018 to January 2019, about 5.4% (311 samples) were positive. Among these, 34.4% (107 samples) positive for enterobacteria and 65.6% (204 samples) for other species. A higher incidence of Klebsiella pneumoniae (34.6% of cases) was observed, followed by Klebsiella sp. (28.9%) and Escherichia coli (26.2%), however, three hemoculture isolates of the species Klebsiella pneumoniae, which were resistant to the three antibiotics (ertapenem, meropenem and imipenem). Two (2) isolates of Enterobacter cloaceae also obtained resistance to the antibiotics used and one (1) species of Enterobacter aerogenes proved resistant only to ertapenem and meropenem and sensitivity to imipenem.

As Enterobacteriaceae são bactérias Gram-negativas e frequentes causadoras de infecções hospitalares. Os carbapenêmicos (CROs) são considerados as mais recentes linhas de defesa contra infecções por microrganismos multirresistentes, no entanto o desenvolvimento de bactérias multirresistentes a esta classe medicamentosa tem prejudicado o tratamento farmacológico. Nesse sentido, o presente estudo trata-se de uma análise retrospectiva, tendo como objetivo avaliar o perfil de resistência e susceptibilidade de bacilos Gram-negativos fermentadores da glicose aos carbapenêmicos em isolados de hemoculturas positivas no período de janeiro de 2018 a janeiro 2019. Das 5.733 hemoculturas realizadas nesse período, cerca de 5,4% (311 amostras) foram positivas. Dentre essas, 34,4% (107 amostras) positivas para enterobactérias e 65,6% (204 amostras) positivas para outras espécies. Foi observada uma maior incidência de Klebsiella pneumoniae (34,6% dos casos), seguido pela Klebsiella sp. (28,9%) e a Escherichia coli (26,2%). Contudo, três (03) isolados de hemocultura da espécie Klebsiella pneumoniae se mostraram resistentes aos três antibióticos (ertapenem, meropenem e imipenem). Dois isolados de Enterobacter cloaceae também obtiveram resistência aos antibióticos utilizados e uma espécie de Enterobacter aerogenes mostrou-se resistente apenas ao ertapenem e meropenem e sensibilidade ao imipenem.

Prevalence of Carbapenem-Resistance among Enterobacteriaceae Isolated from Patients Attending Uduth Sokoto, North West Nigeria

Themajorproblemthreateningthecontinuedsuccessofantimicrobialdrugsisthedevelopmentofresistantorganisms.Thisstudywasdesignedtodeterminetheoccurrenceandprevalenceofcarbapenemresistanceamongenterobacteriaceaeisolatedfromin-patientsattendingUsmanuDanfodiyoUniversityTeachingHospital(UDUTH)Sokoto,andalsotodeterminetheantimicrobialsusceptibilitypatternsoftheorganismsisolated.TheMethodologyinvolvestheuseoftrypticasesoybrothcontaining10μgofcarbapenem(imipenem)forprimaryisolation.ThesecondaryisolationinvolvestheuseofMacConkeyagarandbiochemical(SimmoncitrateAgar)foridentification,andthenantimicrobialsusceptibilitytestingbythediskdiffusionmethod.Atotalof191stoolsamplesfrommaleandfemalein-patientswithintherangeof2-60yearswerescreenedforthe gastrointestinalcolonizationofCarbapenemResistanceEnterobacteriacea(CRE).DatawereanalysedusingStatisticalPackageforSocialSciences(SPSS)windowsversion21.Theoverallprevalencewas15.7%Theprevalenceratewasfoundexclusivelyinmales.Thehighestprevalencewasfoundwithintheagegroupof30-40years.Escherichiacoli,Klebsiellaspecies,Pseudomonasspeciesandsalmonellaspecieswerethebacteriaisolated.ThemostpredominantbacteriaisolatedwereEscherichiacoli,accountingfor85(9.4%)resistantstrains,Klebsiellaspeciesaccountingfor50(8%)resistantstrain,Pseudomonasaeruginosa21%andSalmonellaspeciesaccountingfor42(0%)noresistantstrainisolated.Themisuseofantibioticsisamajorfactorresponsibleforthehighrateofbacterialresistance.Improvementonthemanagementandpersonalhygiene,aswellastheappropriateuseofantibioticswouldreducetheprevalenceofNosocomialCarbapenemResistance(NCR)especiallyamongprolongedhospitalizedpatients

Antibiotic resistance and virulence factors among Escherichia coli isolates from avian organic fertilizer / Resistência a antibióticos e fatores de virulência entre isolados de Escherichia coli provenientes de fertilizante orgânico aviário

ABSTRACT: Brazilian poultry industry generates large amounts of organic waste, such as chicken litter, which is often used in agriculture. Among the bacteria present in organic fertilizer are members of the Enterobacteriaceae family, such as Escherichia coli. The aim of this study was to analyze the profile of virulence factors and antimicrobial resistance of E. coli isolates from avian organic fertilizer. A total of 47 E. coli strains were tested through Polymerase chain reaction to detect virulence genes (hlyF, iss, ompT, iutA and iroN). Fourteen antimicrobials were used to test antimicrobial susceptibility in the strains. Genes characteristic of Avian Pathogenic E. coli (APEC) were reported among the strains, with the hlyF, iss and ompT genes being the most prevalent. The isolates showed high resistance (˃50%) to tetracycline, gentamicin, cefotaxime, nitrofurantoin, trimethoprim-sulfamethoxazole and ampicillin. Multidrug resistance was reported in a great number of strains (>70%). The results showed the presence of APEC cells with virulence genes and antimicrobial resistance after 15 days of the windrowing process in poultry houses, it means this process should be improved to eliminate these cells.

RESUMO: A indústria avícola brasileira gera grandes quantidades de resíduos orgânicos, como a cama de frango, utilizada frequentemente na agricultura. Entre as bactérias presentes neste fertilizante orgânico estão os membros da família Enterobacteriaceae, entre eles a Escherichia coli. O objetivo deste estudo foi analisar o perfil de fatores de virulência e resistência antimicrobiana de isolados de E. coli provenientes do fertilizante orgânico aviário. Um total de 47 cepas de E. coli foram testadas por meio da reação em cadeia da polimerase para detectar genes de virulência (hlyF, iss, ompT, iutA e iroN). Quatorze antimicrobianos foram utilizados para testar a susceptibilidade antimicrobiana nos isolados. Genes característicos de E. coli Patogênica Aviária (APEC) foram encontrados entre os isolados, sendo os genes hlyF, iss e ompT os mais prevalentes. Os isolados apresentaram alta resistência (˃50%) à tetraciclina, gentamicina, cefotaxima, nitrofurantoína, trimetoprima-sulfametoxazole e ampicilina. Múltipla resistência a drogas antimicrobianas foi encontrada em grande número de isolados (>70%). Os resultados obtidos mostraram a presença de células APEC portando genes de virulência e resistência a antimicrobianos após 15 dias de processo de empilhamento nas granjas, indicando que o processo necessita de um aperfeiçoamento para eliminar estas células.

Characterisation of plasmid-mediated rmtB-1 in Enterobacteriaceae clinical isolates from São Paulo, Brazil

OBJECTIVES The emergence of 16S rRNA methyltranferases (16 RMTAses) has jeopardised the clinical use of aminoglycosides. RmtB is one of the most frequently reported in Gram-negatives worldwide. In this study, we aimed to estimate the frequency of 16S RMTAses encoding genes in Enterobacteriaceae isolated in a three-month period from a tertiary Brazilian hospital. METHODS All Gram-negatives classified as resistant to amikacin, gentamicin, and tobramycin by agar screening were selected for analysis. The presence of 16SRMTases encoding genes was verified by polymerase chain reaction (PCR). Antimicrobial susceptible profile was determined by broth microdilution. The genetic relationship among these isolates was accessed by pulsed field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Selected RmtB-producing isolates were characterised by whole genome sequencing (WGS) analysis. RESULTS Twenty-two of 1,052 (2.1%) Enterobacteriaceae were detected as producers of RmtB-1 [Klebsiella pneumoniae (n = 21) and Proteus mirabilis (n = 1)]. blaKPC-2 was identified among 20 RmtB-1-producing K. pneumoniae isolates that exhibited an identical PFGE and MLST (ST258) patterns. Two K. pneumoniae isolates, the A64216 (not harboring bla KPC-2), A64477 (harboring bla KPC-2) and one P. mirabilis isolate (A64421) were selected for WGS. rmtB-1 and bla KPC-2 genes were carried by distinct plasmids. While a plasmid belonging to the IncFIIk group harbored rmtB-1 in K. pneumoniae, this gene was carried by a non-typable plasmid in P. mirabilis. In the three analysed plasmids, rmtB-1 was inserted on a transposon, downstream a Tn2. CONCLUSION Our findings suggested that the rmtB-1 was harbored by plasmids distinct from those previously reported in Bolivia and China. It suggests that multiple mobilization events might have occurred in South America.

Inactivación del carbapenémico, un método alternativo para detectar carbapenemasa tipo KPC en Enterobacteriaceae / Carbapenem inactivation, an alternative method to detect carbapenemase type KPC in Enterobacteriacea

Objetivo: Evaluar al método de inactivación del carbapenémico (MIC*) frente a técnicas como el Test de Hodge modificado (THM), ácido 3-aminofenilborónico (APB) y la reacción en cadena de la polimerasa en enterobacterias productoras de carbapenemasas (EPC) tipo KPC. Materiales y métodos: Se seleccionaron 88 aislados clínicos de K. pneumoniae, K. oxytoca, E.coli, S. marcescens, C. freundii sensibles y 91 resistentes a los carbapenémicos. El APB y el método MIC* se realizaron siguiendo las publicaciones originales. El THM se realizó de acuerdo al CLSI 100S Edición 26-2016. El gen blaKPC se identificó por multiplex PCR. Resultados: El MIC* en EPC tipo KPC presentó una sensibilidad/especificidad cercana al 100% y kappa de 1 comparado con la PCR; se observó la ausencia de halo en todas los aislados EPC tipo KPC a diferencia de los aislados sensibles a los cabapenémicos que presentaron halo > 19mm. Se observó el 3 % de resultados falsos positivos y el 5 % de falsos negativos en THM y ABP respectivamente. Discusión y conclusiones: El MIC* y la PCR demuestran superioridad al THM y ABP para identificar carbapenemasas tipo KPC en EPC. Se recomienda su uso de forma rutinaria dentro del algoritmo para la contención de infecciones por este tipo de patógenos.

Objective: To compare the carbapenem inactivation method (CIM *) with the Modified Hodge Test (MHT), the acid 3-aminophenylboronic test(APB) and the polymerase chain reaction (PCR) detection of the blaKPC gene for the identification of KPC carbapenemase producing Enterobacteriaceae (ECP). Materials and Methods: We selected 88 susceptible and 91 carbapenems resistant clinical isolates of Klebsiella pneumoniae, Klebsiella oxytoca, Escherichia coli, Serratia marcescens and Citrobacter freundii. We performed APB and CIM* according to previously published methods and the MHT according to CLSI 100S Edition 26-2016. The blaKPC gene was identified by PCR multiplex. Results: The CIM* had a sensitivity and specificity close to 100% and a kappa score of 1 compared with gold standard PCR. The absence of zone diameter was observed in all isolated KPC producers, unlike in isolates susceptible to carbapenems, where a zone diameter >19mm was observed. Three percent of false positive and five percent of false negative was observed in THM and ABP respectively. Discussion and conclusions: The CIM* and the PCR were better than MHT and ABP at identifying carbapenemases in ECP. We recommend the routine use of the CIM* within the algorithm for ECP infection control.

Comparison of double disc synergy test and phenotypic confirmatory disc diffusion test for detection of ESBL production and their antimicrobial resistance pattern.

Extended spectrum beta- lactamases (ESBLs) continue to be a major problem in clinical setup worldwide. An attempt is made to detect ESBL production among Enterobacteriaceae members by phenotypic methods, which is easier to perform in all laboratories. A total of 138 multi-drug resistant strains from pyogenic infection were tested for ESBL production by Double disc synergy test (DDST) and Phenotypic confirmatory disc diffusion test (PCDDT). Of the 84 ESBL producer identified, PCDDT detects 71 (84.5%) whereas DDST detects 52 (61.9%) as ESBL producers. Continued detection of ESBL is essential for proper disease management. PCDDT is better and easy test for screening than DDST. Confirmation has to be done by molecular methods.

Próteses totais removíveis como reservatório de microrganismos oportunistas / Removable total prostheses as reservoir of oportunists microrganisms

Este estudo avaliou a ocorrência de leveduras do gênero Candida sp além dos principais microrganismos periodontopatogênicos e enterobactérias na saliva, em mucosa e no biofilme aderido à prótese total, correlacionando com aspectos clínicos e condição de higiene bucal de 90 indivíduos edêntulos e portadores de prótese total, por meio de métodos moleculares (PCR). Espécimes clínicos intrabucais foram coletados desses indivíduos após avaliação das condições sócio-econômicas e comportamentais. A microbiota bucal dos pacientes foi caracterizada por meio da obtenção de amostras de biofilme aderido às próteses totais, mucosa e saliva, as quais foram processadas, por meio de PCR. As inter-relações entre os diferentes microrganismos foram determinadas por meio dos testes de Qui-quadrado e Mann- Whitney. Verificaram-se diferenças na ocorrência de Prevotella intermedia e Enterobacteriaceae na saliva dos pacientes edêntulos, o mesmo ocorrendo com Enterobacteriaceae, Camphylobacter rectus e gênero Pseudomonas no biofilme aderido às próteses totais. As condições de higiene bucal e estado de conservação da prótese total precários favoreceram a ocorrência de leveduras do tipo Candida sp, em especial Candida albicans, em níveis estatisticamente significante nas amostras de mucosa e biofilme aderido à prótese total, tornando este dispositivo protético um potencial reservatório de leveduras e bactérias entéricas que podem ser de relevância na patogênese das infecções oportunistas

The aim of this study was to evaluate the occurrence of major periodontopathogenic microorganisms and Enterobacteriaceae and biofilm adhered to the denture, mucosa and saliva in 90 edentulous subjects with complete dentures, using molecular methods (PCR). Clinical specimens were collected from these individuals after assessing the socio-economic circumstances and behavioral. The oral microbiota of patients was characterized by obtaining samples of the biofilm adhered to the dental prothesis and saliva, for detection of major pathogens using PCR. The possibility of inter-relationships between different microorganisms was determined using the chi-square test and Mann-Whitney test. There were differences in the occurrence of Prevotella intermedia and Enterobacteriaceae in the saliva of edentulous patients, likewise, Enterobacteriaceae, Pseudomonas, Camphylobacter rectus and the biofilm attached to denture patients. The conditions for oral hygiene and stat of preservation of prosthesis total precarious favored the occurrence of yeasts of the Candida sp, particularly Candida albicans, statistically significant levels in samples of mucosa and biofilm acceded to total prosthesis, prothetic device, making it a potential reservoir of enteric bacteria and yeasts that may be of relevance in the pathogenesis of opportunistic infections

Próteses totais removíveis como reservatório de microrganismos oportunistas / Removable total prostheses as reservoir of oportunists microrganisms

Este estudo avaliou a ocorrência de leveduras do gênero Candida sp além dos principais microrganismos periodontopatogênicos e enterobactérias na saliva, em mucosa e no biofilme aderido à prótese total, correlacionando com aspectos clínicos e condição de higiene bucal de 90 indivíduos edêntulos e portadores de prótese total, por meio de métodos moleculares (PCR). Espécimes clínicos intrabucais foram coletados desses indivíduos após avaliação das condições sócio-econômicas e comportamentais. A microbiota bucal dos pacientes foi caracterizada por meio da obtenção de amostras de biofilme aderido às próteses totais, mucosa e saliva, as quais foram processadas, por meio de PCR. As inter-relações entre os diferentes microrganismos foram determinadas por meio dos testes de Qui-quadrado e Mann- Whitney. Verificaram-se diferenças na ocorrência de Prevotella intermedia e Enterobacteriaceae na saliva dos pacientes edêntulos, o mesmo ocorrendo com Enterobacteriaceae, Camphylobacter rectus e gênero Pseudomonas no biofilme aderido às próteses totais. As condições de higiene bucal e estado de conservação da prótese total precários favoreceram a ocorrência de leveduras do tipo Candida sp, em especial Candida albicans, em níveis estatisticamente significante nas amostras de mucosa e biofilme aderido à prótese total, tornando este dispositivo protético um potencial reservatório de leveduras e bactérias entéricas que podem ser de relevância na patogênese das infecções oportunistas

The aim of this study was to evaluate the occurrence of major periodontopathogenic microorganisms and Enterobacteriaceae and biofilm adhered to the denture, mucosa and saliva in 90 edentulous subjects with complete dentures, using molecular methods (PCR). Clinical specimens were collected from these individuals after assessing the socio-economic circumstances and behavioral. The oral microbiota of patients was characterized by obtaining samples of the biofilm adhered to the dental prothesis and saliva, for detection of major pathogens using PCR. The possibility of inter-relationships between different microorganisms was determined using the chi-square test and Mann-Whitney test. There were differences in the occurrence of Prevotella intermedia and Enterobacteriaceae in the saliva of edentulous patients, likewise, Enterobacteriaceae, Pseudomonas, Camphylobacter rectus and the biofilm attached to denture patients. The conditions for oral hygiene and stat of preservation of prosthesis total precarious favored the occurrence of yeasts of the Candida sp, particularly Candida albicans, statistically significant levels in samples of mucosa and biofilm acceded to total prosthesis, prothetic device, making it a potential reservoir of enteric bacteria and yeasts that may be of relevance in the pathogenesis of opportunistic infections

Diagnóstico microbiológico e histopatológico de mortalidade em avestruzes (Struthio camelus) / Microbiological and histological diagnosis in mortality of ostrich (Struthio camelus)

Several young ostrich, including nestlings, with lassitude and inappetence followed by death or victim of sudden death were immediately brought to diagnosis at an Animal Health Laboratory. At necropsy, animals presented hemorrhage and altered content of the vitelline sac, and necrotic foci in the small intestine; one animal showed necrotic pleuropneumonia with psammomatosus bodies in the lung parenchyma. The cultures from different samples revealed Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Serratia marcescens, Enterobacter aglomerans, and Pseudomonas mendocina. It was suggested one case of septicemia in an animal with exclusive growth of K. pneumoniae isolated from samples of small intestine, lung, and liver.