Intracellular proliferation of Legionella pneumophila in Hartmannella vermiformis in aquatic biofilms grown on plasticized polyvinyl chloride

The need for protozoa for the proliferation of Legionella pneumophila in aquatic habitats is still not fully understood and is even questioned by some investigators. This study shows the in vivo growth of L. pneumophila in protozoa in aquatic biofilms developing at high concentrations on plasticized polyvinyl chloride in a batch system with autoclaved tap water. The inoculum, a mixed microbial community including indigenous L. pneumophila originating from a tap water system, was added in an unfiltered as well as filtered (cellulose nitrate, 3.0-mum pore size) state. Both the attached and suspended biomasses were examined for their total amounts of ATP, for culturable L. pneumophila, and for their concentrations of protozoa. L. pneumophila grew to high numbers (6.3 log CFU/cm(2)) only in flasks with an unfiltered inoculum. Filtration obviously removed the growth-supporting factor, but it did not affect biofilm formation, as determined by measuring ATP. Cultivation, direct counting, and 18S ribosomal DNA-targeted PCR with subsequent sequencing revealed the presence of Hartmannella vermiformis in all flasks in which L. pneumophila multiplied and also when cyclobeximide had been added. Fluorescent in situ hybridization clearly demonstrated the intracellular growth of L. pneumophila in trophozoites of H. vermiformis, with 25.9% +/- 10.5% of the trophozoites containing L. pneumophila on day 10 and >90% containing L. pneumophila on day 14. Calculations confirmed that intracellular growth was most likely the only way for L. pneumophila to proliferate within the biofilm. Higher biofilm concentrations, measured as amounts of ATP, gave higher L. pneumophila concentrations, and therefore the growth of L. pneumophila within engineered water systems can be limited by controlling biofilm formation

Polaromonas and Hydrogenophaga species are the predominant bacteria cultured from granular activated carbon filters in water treatment

AIM: Identification of the predominating cultivable bacteria in granular activated carbon (GAC) filters used in a variety of water treatment plants for selecting representative strains to study the role of bacteria in the removal of dissolved organic matter. METHODS AND RESULTS: Bacterial isolates were collected from 21 GAC filters in nine water treatment plants treating either ground water or surface water with or without oxidative pretreatment. Enrichment of samples in dilute liquid medium improved culturability of the bacteria by approximately log unit, to 9% up to 70% of the total cell counts. Genomic fingerprinting and 16S rDNA sequence analysis revealed that most (68%) of the isolates belonged to the Betaproteobacteria and 25% were identified as Alphaproteobacteria. The number of different genera within the Betaproteobacteria was higher in the GAC filters treating ozonated water than in the filters treating nonozonated water. Polaromonas was observed in nearly all of the GAC filters (86%), and the genera Hydrogenophaga, Sphingomonas and Afipia were observed in 43%, 33% and 29% of the filter beds, respectively. AFLP analysis revealed that the predominating genus Polaromonas included a total of 23 different genotypes. CONCLUSIONS: This study is the first to demonstrate that Polaromonas, which has mainly been observed in ultraoligotrophic freshwater environments, is a common component of the microbial community in GAC filters used in water treatment. SIGNIFICANCE AND IMPACT OF THE STUDY: The predominance of ultraoligotrophic bacteria in the GAC filters indicates that very low concentrations of substrates are available for microbial growth. Polaromonas species are suited for further studies on the nutritional versatility and growth kinetics enabling the modelling of biodegradation processes in GAC filter

Ionophore resistance and potential risk of ionophore driven co-selection of clinically relevant antimicrobial resistance in poultry

Today’s intensive broiler production is highly dependent on ionophore coccidiostats. These antiparasitic agents are applied as feed additives. The most important group, the ionophores, not only exhibit antiparasitic activity but are also antimicrobial against Gram-positive bacteria. Because these ionophores are not used in humans, it is widely assumed their use does not impact human health. However, recent Swedish and Norwegian research shows that ionophores can cause the co-selection of vancomycin resistance in enterococci. Vancomycin is a critically important antimicrobial for human medicine. Because of its limited relevance for human medicine, monitoring of antimicrobial resistance against ionophores, in particular salinomycin (SAL), was discontinued several years ago. From historical data, however, it can be retrieved that a fair percentage of the Enterococcus faecium and Enterococcus faecalis isolates from poultry origin show SAL resistance. The current research was initiated to determine whether ionophore resistance in enterococci from poultry co-occurs with other clinically relevant types of antimicrobial resistance. A set of 137 E. faecium and E. faecalis isolates from poultry products from conventional and organic origin was phenotypically characterized. The resistance profiles were statistically analysed and subsequently a selection of 20 isolates was DNA sequenced and analysed for the presence of resistance genes. Almost 40% of the isolates appeared to have a SAL resistant phenotype. Results of the statistical analysis showed a significant correlation between the presence of SAL resistance and erythromycin (ERY) resistance in E. faecium from conventional farming. The same correlation was also observed in E. faecalis from both conventional and organic origin. Besides this, both E. faecium and E. faecalis show significant correlation between the presence of SAL and tetracycline (TET) resistance and SAL and ampicillin (AMP) resistance. DNA sequencing results show good agreement between the observed phenotypic resistance and the identified resistance genes. In all of the isolates showing phenotypic SAL resistance the narAB genes could be identified. Moreover, in most isolates narAB is found to be located on a plasmid carrying additional genes for other types of antibiotic resistance. This is an alarming observation, since it implies that the use of ionophores may drive the transfer and dissemination of other, clinically relevant types of antimicrobial resistance by co-selection. These results question the sustainability of the prophylactic use of ionophores in broiler production. More research is needed to determine the extent and the impact of this issue. It is unknown whether ionophore resistance also occurs in other Gram-positive bacteria, like Staphylococcus aureus. It is plausible that ionophore resistance genes other than narAB will be present in poultry associated bacteria. E. faecium and E. faecalis are an important cause of hospital-acquired infections. Analysis of human enterococcal isolates should reveal whether transmission from the poultry reservoir is occurring and thus whether the use of ionophores in poultry impacts human healthcare

Intracellular proliferation of Legionella pneumophila in Hartmannella vermiformis in aquatic biofilms grown on plasticized polyvinyl chloride

The need for protozoa for the proliferation of Legionella pneumophila in aquatic habitats is still not fully understood and is even questioned by some investigators. This study shows the in vivo growth of L. pneumophila in protozoa in aquatic biofilms developing at high concentrations on plasticized polyvinyl chloride in a batch system with autoclaved tap water. The inoculum, a mixed microbial community including indigenous L. pneumophila originating from a tap water system, was added in an unfiltered as well as filtered (cellulose nitrate, 3.0-mum pore size) state. Both the attached and suspended biomasses were examined for their total amounts of ATP, for culturable L. pneumophila, and for their concentrations of protozoa. L. pneumophila grew to high numbers (6.3 log CFU/cm(2)) only in flasks with an unfiltered inoculum. Filtration obviously removed the growth-supporting factor, but it did not affect biofilm formation, as determined by measuring ATP. Cultivation, direct counting, and 18S ribosomal DNA-targeted PCR with subsequent sequencing revealed the presence of Hartmannella vermiformis in all flasks in which L. pneumophila multiplied and also when cyclobeximide had been added. Fluorescent in situ hybridization clearly demonstrated the intracellular growth of L. pneumophila in trophozoites of H. vermiformis, with 25.9% +/- 10.5% of the trophozoites containing L. pneumophila on day 10 and >90% containing L. pneumophila on day 14. Calculations confirmed that intracellular growth was most likely the only way for L. pneumophila to proliferate within the biofilm. Higher biofilm concentrations, measured as amounts of ATP, gave higher L. pneumophila concentrations, and therefore the growth of L. pneumophila within engineered water systems can be limited by controlling biofilm formation

Surveillance en monitoring van Salmonella bij de mens, landbouwhuisdieren en in voedsel in Nederland, 2019

Na een jarenlange continu dalende trend in Nederland van Salmonella-infecties bij de mens is het aantal sinds 2013 gestabiliseerd op ruim 1500 laboratoriumbevestigde patiënten per jaar met een geschatte 26.000 patiënten in de gehele bevolking. Deze stabilisatie komt overeen met het beeld dat wordt gezien in de monitoring van landbouwhuisdieren en levensmiddelen. Opvallend in 2019 was de stijging in humane gevallen met S. Enteritidis-infecties (meestal gerelateerd aan eieren) en de daling van infecties met S. Typhimurium (meestal gerelateerd aan varkensvlees). Tevens is vanaf 2012 een significante stijging te zien van het aandeel invasieve infecties ten opzichte van het totaal aantal infecties door Salmonella. Alhoewel er in 2019 6 clusters werden gedetecteerd, was het een relatief rustig jaar wat betreft uitbraken (de laatste grote uitbraak dateert van 2016/2017 met S. Enteritidis in Poolse eieren). Whole genome sequencing (WGS) speelt een steeds belangrijkere rol in de surveillance bij het aantonen van clusters en het relateren van bronnen aan uitbraken. Alhoewel redelijk stabiel, is de resistentie tegen fluoroquinolonen nog steeds een zorg, evenals multiresistentie en ESBL(Extended Spectrum Beta Lactamases)-producerende Salmonella-isolaten