Sanitary importance of arcobacter

The genus Arcobacter, belongs to the family Campylobacteraceae, along with. Some species, are considered emerging pathogens and have been associated with gastrointestinal diseases and bacteraemia in humans and with diarrhoea, mastitis and abortions in animals. Diarrhoea, is the most common clinical presentation in humans. However, still are not fully defined the routes of transmission, the virulence factors present in pathogenic strains have not been determined and the isolation and identification methods are still deficient. All of these factors contribute to a possible underestimation of the sanitary importance of this genus. In this thesis it has been demonstrated the existence of five new species of this genus. In their differentiation, new tools have been used, such as the Matrix Assisted Laser Desorption Ionization Time of Flight (MALDI TOF) Multilocus Phylogenetic Analysis (MLPA) with concatenated 5 genes (rpoB, gyrB, hsp60, gyrA and atpA). Furthermore, the performance of 5 molecular identification methods was compared and none of them was useful for all the strains studied. In this regard, the updating of 16S rRNARFLP method for the identification of all species proved very useful. Moreover, in this thesis was a great diversity of species of this genus in samples of shellfish and sewage. Both matrices have epidemiological importance; however, they have been poorly studied. It was also demonstrated that the use in parallel of direct culturing and post enrichment, as well as the incubation under aerobic and microaerophilic, enhances the recovery and diversity of arcobacters. Regarding the virulence of Arcobacter spp., most of them proved to be potential enteropathogens for man, as they showed the presence of virulence factors and were able to adhere and invade human intestinal cells (Caco-2). Finally, it was demonstrated that Arcobacter may be confused with Campylobacter sp., generating an underestimation of their sanitary importance.El género Arcobacter incluye especies consideradas patógenos emergentes, ya que se han asociado con patologías gastrointestinales y bacteriemia en humanos y con diarrea, mastitis y abortos en animales. Sin embargo, aún no se ha definido completamente las rutas de transmisión, los factores de virulencia presentes en cepas patogénicas no se han determinado y los métodos de aislamiento e identificación aún son deficientes. Todo esto genera una subestimación de la importancia sanitaria de Arcobacter. En esta tesis doctoral se demostró la existencia de 5 nuevas especies de este género. Para su diferenciación se utilizó herramientas nuevas, tales como el Matrix Assisted Laser desorption Ionization Time of Flight (MALDI TOF) y un Multilocus Phylogenetic Analysis (MLPA) con 5 genes concatenados (rpoB, gyrB, hsp60, atpA y gyrA). Considerando el número de especies incluidas actualmente en el género (n=17), se evaluó si 5 de los métodos moleculares de identificación son todavía útiles para la identificación de las especies para las que se habían definido o si se generan confusiones, demostrrando que todos ellos generan algún error que osciló entre el 16,8% y 67,4% de las cepas estudiadas. En este sentido, la ampliación del método 16S rRNA-RFLP para poder identificar todas las especies resultó ser de gran utilidad. Por otra parte, en esta tesis se observó una gran diversidad de especies de este género en muestras de mariscos y aguas residuales. Ambas matrices han sido poco estudiados a pesar de su importancia epidemiológica. Más aún, se demostró que el uso en paralelo del cultivo por siembra directa y post enriquecimiento, además de incubación en aerobiosis y microaerofilia, favorece la recuperación de una mayor diversidad. Por otra parte, también se demostró que la mayoría de las especies de este género, en especial algunas cepas A. butzleri, A. cryaerophilus, A. skirrowii, A. trophiarum y A. defluvii, son potenciales enteropatógenos para el hombre, ya que presentaron factores de virulencia y fueron capaces de adherir e invadir células intestinales humanas (Caco-2). Por último, se demostró que Arcobacter puede ser confundido con Campylobacter sp., lo que puede contribuir aún más a subestimar su importancia sanitaria

SIDEROPHORES FOR SELECTIVE SOLID PHASE EXTRACTION OF STRATEGIC ELEMENTS

All over the world, industrial mining is leaving contaminated areas and dumps that, although being full of valuable metals, have high concentrations of toxic heavy metals that pollute the environment. The development of sustainable alternative biomining and bioremediation processes offers the potential to fully exploit these unexploited mining sites

Cronobacter condimenti sp. nov., isolated from spiced meat, and Cronobacter universalis sp. nov., a species designation for Cronobacter sp. genomospecies 1, recovered from a leg infection, water, and food ingredients

A re-evaluation of the taxonomic position of five strains, one assigned to Cronobacter sakazakii (strain 1330T), two previously assigned to Cronobacter genomospecies 1 (strains NCTC 9529T and 731) and two as Cronobacter turicensis (strains 96 and 1435) was carried out. The analysis included a phenotypic characterization, sequencing of the 16S rRNA gene and multilocus sequence analysis (MLSA) of seven housekeeping genes (atpD, fusA, glnS, gltB, gyrB, infB, ppsA; 3036 bp). The 16S rRNA gene sequence analysis and MLSA showed strain 1330T, isolated from spiced meat purchased in Slovakia, to form an independent phylogenetic line. Cronobacter dublinensis was the closest neighbour species on the basis of the MLSA. DNA–DNA reassociation and phenotypic analysis revealed that strain 1330T represented a novel species, for which the name Cronobacter condimenti sp. nov. is proposed, type strain 1330T = CECT 7863T, = LMG 26250T). The four bacterial strains NCTC 9529T, 731, 96 and 1435, isolated from water, a leg infectionand two food ingredients; onion powder and rye flour, repectively, showed on the phylogenetic tree to cluster together within an independent phylogenetic line, with Cronobacter turicensis as the closest species. The DNA–DNA hybridization data and the phenotypic characterization confirmed that these strains represented a novel species, for which the name Cronobacter universalis sp. nov. is proposed with type strain NCTC 9529T = CECT 7864T, = LMG 26249T

Head-to-head comparison of CAMPYAIR aerobic culture medium versus standard microaerophilic culture for Campylobacter isolation from clinical samples

Campylobacter spp. are considered the most frequent cause of acute gastroenteritis worldwide. However, outside high-income countries, its burden is poorly understood. Limited published data suggest that Campylobacter prevalence in low- and middle-income countries is high, but their reservoirs and age distribution are different. Culturing Campylobacter is expensive due to laboratory equipment and supplies needed to grow the bacterium (e.g., selective culture media, microaerophilic atmosphere, and a 42°C incubator). These requirements limit the diagnostic capacity of clinical laboratories in many resource-poor regions, leading to significant underdiagnosis and underreporting of isolation of the pathogen. CAMPYAIR, a newly developed selective differential medium, permits Campylobacter isolation without the need for microaerophilic incubation. The medium is supplemented with antibiotics to allow Campylobacter isolation in complex matrices such as human feces. The present study aims to evaluate the ability of the medium to recover Campylobacter from routine clinical samples. A total of 191 human stool samples were used to compare the ability of CAMPYAIR (aerobic incubation) and a commercial Campylobacter medium (CASA, microaerophilic incubation) to recover Campylobacter. All Campylobacter isolates were then identified by MALDI-TOF MS. CAMPYAIR showed sensitivity and specificity values of 87.5% (95% CI 47.4%–99.7%) and 100% (95% CI 98%–100%), respectively. The positive predictive value of CAMPYAIR was 100% and its negative predictive value was 99.5% (95% CI 96.7%–99.9%); Kappa Cohen coefficient was 0.93 (95% CI 0.79–1.0). The high diagnostic performance and low technical requirements of the CAMPYAIR medium could permit Campylobacter culture in countries with limited resources

CAMPYAIR, a New Selective, Differential Medium for Campylobacter spp. Isolation without the Need for Microaerobic Atmosphere

Campylobacter spp. are considered the most frequent bacterial cause of acute gastroenteritis worldwide. Although the diarrhea produced by these bacteria is self-limiting, the pathogen has been associated with severe long-term sequelae following acute signs and symptoms of the illness. However, research on Campylobacter is hampered by costs and technical requirements for isolating and culturing the bacterium, especially in low and middle-income countries. Therefore, attempts have been made to simplify these culture methods and to reduce costs associated with conducting research on Campylobacter. Recently, a liquid medium which allows selective enrichment of Campylobacter using aerobic incubation has been described. However, a solid medium is also needed for the isolation of pure colonies, enumeration of bacterial populations, and other studies on the pathogen. Therefore, a new medium (CAMPYAIR) was developed, based on the formulation of the liquid medium. CAMPYAIR is a solid chromogenic medium that supports the growth of Campylobacter isolates within 48 h of incubation in aerobic atmospheres. Moreover, CAMPYAIR contains antibiotic supplements with an enhanced ability to recover Campylobacter from environmental samples that may also contain non-campylobacter bacteria. The addition of the indicator 2,3,5-triphenyltetrazolium (TTC) to the medium differentiates Campylobacter from other bacteria growing on the media. The findings from studies on CAMPYAIR suggest that the utilization of the new selective, differential medium could help to reduce the costs, equipment, and technical training required for Campylobacter isolation from clinical and environmental samples

CAMPYAIR, a New Selective, Differential Medium for <i>Campylobacter</i> spp. Isolation without the Need for Microaerobic Atmosphere

Campylobacter spp. are considered the most frequent bacterial cause of acute gastroenteritis worldwide. Although the diarrhea produced by these bacteria is self-limiting, the pathogen has been associated with severe long-term sequelae following acute signs and symptoms of the illness. However, research on Campylobacter is hampered by costs and technical requirements for isolating and culturing the bacterium, especially in low and middle-income countries. Therefore, attempts have been made to simplify these culture methods and to reduce costs associated with conducting research on Campylobacter. Recently, a liquid medium which allows selective enrichment of Campylobacter using aerobic incubation has been described. However, a solid medium is also needed for the isolation of pure colonies, enumeration of bacterial populations, and other studies on the pathogen. Therefore, a new medium (CAMPYAIR) was developed, based on the formulation of the liquid medium. CAMPYAIR is a solid chromogenic medium that supports the growth of Campylobacter isolates within 48 h of incubation in aerobic atmospheres. Moreover, CAMPYAIR contains antibiotic supplements with an enhanced ability to recover Campylobacter from environmental samples that may also contain non-campylobacter bacteria. The addition of the indicator 2,3,5-triphenyltetrazolium (TTC) to the medium differentiates Campylobacter from other bacteria growing on the media. The findings from studies on CAMPYAIR suggest that the utilization of the new selective, differential medium could help to reduce the costs, equipment, and technical training required for Campylobacter isolation from clinical and environmental samples

Updated 16S rRNA-RFLP method for the identification of all currently characterised <it>Arcobacter</it> spp

Abstract Background Arcobacter spp. (family Campylobacteraceae) are ubiquitous zoonotic bacteria that are being increasingly recognised as a threat to human health. A previously published 16S rRNA-RFLP Arcobacter spp. identification method produced specific RFLP patterns for the six species described at that time, using a single endonuclease (MseI). The number of characterised Arcobacter species has since risen to 17. The aim of the present study was to update the 16S rRNA-RFLP identification method to include all currently characterised species of Arcobacter. Results Digestion of the 16S rRNA gene with the endonuclease MseI produced clear, distinctive patterns for 10 of the 17 species, while the remaining species shared a common or very similar RFLP pattern. Subsequent digestion of the 16S rRNA gene from these species with the endonucleases MnlI and/or BfaI generated species-specific RFLP patterns. Conclusions 16S rRNA-RFLP analysis identified 17 Arcobacter spp. using either polyacrylamide or agarose gel electrophoresis. Microheterogeneities within the 16S rRNA gene, which interfered with the RFLP identification, were also documented for the first time in this genus, particularly in strains of Arcobacter cryaerophilus isolated from animal faeces and aborted foetuses.</p

The Use of Two Culturing Methods in Parallel Reveals a High Prevalence and Diversity of Arcobacter spp. in a Wastewater Treatment Plant

The genus Arcobacter includes species considered emerging food and waterborne pathogens. Despite Arcobacter has been linked to the presence of faecal pollution, few studies have investigated its prevalence in wastewater, and the only isolated species were Arcobacter butzleri and Arcobacter cryaerophilus. This study aimed to establish the prevalence of Arcobacter spp. at a WWTP using in parallel two culturing methods (direct plating and culturing after enrichment) and a direct detection by m-PCR. In addition, the genetic diversity of the isolates was established using the ERIC-PCR genotyping method. Most of the wastewater samples (96.7%) were positive for Arcobacter and a high genetic diversity was observed among the 651 investigated isolates that belonged to 424 different ERIC genotypes. However, only few strains persisted at different dates or sampling points. The use of direct plating in parallel with culturing after enrichment allowed recovering the species A. butzleri, A. cryaerophilus, Arcobacter thereius, Arcobacter defluvii, Arcobacter skirrowii, Arcobacter ellisii, Arcobacter cloacae, and Arcobacter nitrofigilis, most of them isolated for the first time from wastewater. The predominant species was A. butzleri, however, by direct plating predominated A. cryaerophilus. Therefore, the overall predominance of A. butzleri was a bias associated with the use of enrichment

Microbial Communities Associated with Farmed Genypterus chilensis: Detection in Water Prior to Bacterial Outbreaks Using Culturing and High-Throughput Sequencing

The red conger eel (Genypterus chilensis, Guichenot) is a native species included in the Chilean Aquaculture Diversification Program due to high commercial demand. In the context of intensified farming, prior reports link two disease outbreaks with emerging pathogens in the Vibrio and Tenacibaculum genera. However, the roles remain unclear for the bacterial community and each specific bacterium is associated with the rearing environment for healthy specimens. The success of red conger eel farming therefore warrants research into the bacterial composition of aquaculture conditions and the antimicrobial susceptibilities thereof. This study used culturing methods and high-throughput sequencing to describe the bacterial community associated with water in which G. chilensis was farmed. With culturing methods, the predominant genera were Vibrio (21.6%), Pseudolteromonas (15.7%), Aliivibrio (13.7%), and Shewanella (7.8%). Only a few bacterial isolates showed amylase, gelatinase, or lipase activity, and almost all showed inhibition zones to commonly-used antibiotics in aquaculture. By contrast, high-throughput sequencing established Paraperlucidibaca, Colwellia, Polaribacter, Saprospiraceae, and Tenacibaculum as the predominant genera, with Vibrio ranking twenty-seventh in abundance. High-throughput sequencing also established a link between previous outbreaks with increased relative abundances of Vibrio and Tenacibaculum. Therefore, monitoring the presence and abundance of these potential pathogens could be useful in providing prophylactic measures to prevent future outbreaks