Felipe C. Cabello

Expression of Borrelia burgdorferi OspC and DbpA is controlled by a RpoN–RpoS regulatory pathway

RpoS and RpoN are two alternative sigma factors typically associated with general stress responses in bacteria. To date, there has been no experimental evidence that RpoS and RpoN can directly control the expression of one another. Herein, using a combined strategy of gene disruption and genetic complementation targeting rpoN and rpoS in Borrelia burgdorferi strain 297, we describe a regulatory network for B. burgdorferi. In this network, RpoN controls the expression of RpoS, which, in turn, governs the expression of two important membrane lipoproteins, outer surface protein C and decorin-binding protein A, and likely other proteins of B. burgdorferi. Our findings provide a foundation for elucidating further key regulatory networks that potentially impact many aspects of B. burgdorferis parasitic strategy, host range, and virulence expression.

Antimicrobial use in aquaculture re‐examined: its relevance to antimicrobial resistance and to animal and human health

The worldwide growth of aquaculture has been accompanied by a rapid increase in therapeutic and prophylactic usage of antimicrobials including those important in human therapeutics. Approximately 80% of antimicrobials used in aquaculture enter the environment with their activity intact where they select for bacteria whose resistance arises from mutations or more importantly, from mobile genetic elements containing multiple resistance determinants transmissible to other bacteria. Such selection alters biodiversity in aquatic environments and the normal flora of fish and shellfish. The commonality of the mobilome (the total of all mobile genetic elements in a genome) between aquatic and terrestrial bacteria together with the presence of residual antimicrobials, biofilms, and high concentrations of bacteriophages where the aquatic environment may also be contaminated with pathogens of human and animal origin can stimulate exchange of genetic information between aquatic and terrestrial bacteria. Several recently found genetic elements and resistance determinants for quinolones, tetracyclines, and β-lactamases are shared between aquatic bacteria, fish pathogens, and human pathogens, and appear to have originated in aquatic bacteria. Excessive use of antimicrobials in aquaculture can thus potentially negatively impact animal and human health as well as the aquatic environment and should be better assessed and regulated.

Vibrio parahaemolyticus diarrhea, Chile, 1998 and 2004.

Analysis of clinical isolates of Vibrio parahaemolyticus from outbreaks in Chile in the cities of Puerto Montt in 2004 and in Antofagasta in 1998 indicated that 23 of 24 isolates from Puerto Montt and 19 of 20 from Antofagasta belonged to the pandemic clonal complex that emerged in Southeast Asia in 1996.

Antibióticos y acuicultura en Chile: consecuencias para la salud humana y animal

Industrial antibiotic usage in agribusinesses and aquaculture is the force that drives the evolution of antibiotic resistant bacteria that produce human and animal disease in many countries. Several studies have demonstrated that most of the industrial use of antibiotics is unnecessary, and that modernization and hygienic changes can reduce this use of antibiotics without negative economic impact. In Chile, industrial aquaculture of salmon has expanded rapidly in the last 20 years becoming a major export business. The exponential growth of this industry has been accompanied by an unrestricted heavy usage of antibiotics in the aquatic environments of lakes, rivers and the ocean, and its impact is being felt in the emergence of antibiotic-resistant bacteria around aquaculture sites and a decrease in the plancktonic diversity in the same areas. The passage of antibiotic resistance genes from aquatic bacteria to human and animal pathogens has been demonstrated, indicating that industrial use of antibiotics in aquaculture affects negatively the antibiotic therapy of human and animal bacterial infections. The Chilean situation triggers important concerns because it includes the use of fluoroquinolones in aquaculture, that are not biodegradable and are able to remain in the environment for years as well as being still effective in treating human infections. The use of large volumes of a wide spectrum of antibiotics in an aquatic environment heavily contaminated with human and animal pathogens also amplifies the opportunities for gene transfer among bacteria, facilitating the emergence of antibiotic resistance and more pathogenic bacterial recombinants. The detection of residual antibiotics in salmons marketed for human consumption that can modify the normal flora of the population also suggests the need for controls on this antibiotic usage and on the presence of residual antibiotics in aquaculture food products. This important problem of public health demands an active dialogue between government officials responsible for protecting public health, aquaculture industry representatives, politicians, consumers and professionals dealing with these matters.

Salmon Aquaculture and Antimicrobial Resistance in the Marine Environment

Antimicrobials used in salmon aquaculture pass into the marine environment. This could have negative impacts on marine environmental biodiversity, and on terrestrial animal and human health as a result of selection for bacteria containing antimicrobial resistance genes. We therefore measured the numbers of culturable bacteria and antimicrobial-resistant bacteria in marine sediments in the Calbuco Archipelago, Chile, over 12-month period at a salmon aquaculture site approximately 20 m from a salmon farm and at a control site 8 km distant without observable aquaculture activities. Three antimicrobials extensively used in Chilean salmon aquaculture (oxytetracycline, oxolinic acid, and florfenicol) were studied. Although none of these antimicrobials was detected in sediments from either site, traces of flumequine, a fluoroquinolone antimicrobial also widely used in Chile, were present in sediments from both sites during this period. There were significant increases in bacterial numbers and antimicrobial-resistant fractions to oxytetracycline, oxolinic acid, and florfenicol in sediments from the aquaculture site compared to those from the control site. Interestingly, there were similar numbers of presumably plasmid-mediated resistance genes for oxytetracycline, oxolinic acid and florfenicol in unselected marine bacteria isolated from both aquaculture and control sites. These preliminary findings in one location may suggest that the current use of large amounts of antimicrobials in Chilean aquaculture has the potential to select for antimicrobial-resistant bacteria in marine sediments.

Antimicrobial resistance and antimicrobial resistance genes in marine bacteria from salmon aquaculture and non‐aquaculture sites

Antimicrobial resistance (AR) detected by disc diffusion and antimicrobial resistance genes detected by DNA hybridization and polymerase chain reaction with amplicon sequencing were studied in 124 marine bacterial isolates from a Chilean salmon aquaculture site and 76 from a site without aquaculture 8 km distant. Resistance to one or more antimicrobials was present in 81% of the isolates regardless of site. Resistance to tetracycline was most commonly encoded by tetA and tetG; to trimethoprim, by dfrA1, dfrA5 and dfrA12; to sulfamethizole, by sul1 and sul2; to amoxicillin, by blaTEM ; and to streptomycin, by strA-strB. Integron integrase intl1 was detected in 14 sul1-positive isolates, associated with aad9 gene cassettes in two from the aquaculture site. intl2 Integrase was only detected in three dfrA1-positive isolates from the aquaculture site and was not associated with gene cassettes in any. Of nine isolates tested for conjugation, two from the aquaculture site transferred AR determinants to Escherichia coli. High levels of AR in marine sediments from aquaculture and non-aquaculture sites suggest that dispersion of the large amounts of antimicrobials used in Chilean salmon aquaculture has created selective pressure in areas of the marine environment far removed from the initial site of use of these agents.

Aquaculture as yet another environmental gateway to the development and globalisation of antimicrobial resistance

Aquaculture uses hundreds of tonnes of antimicrobials annually to prevent and treat bacterial infection. The passage of these antimicrobials into the aquatic environment selects for resistant bacteria and resistance genes and stimulates bacterial mutation, recombination, and horizontal gene transfer. The potential bridging of aquatic and human pathogen resistomes leads to emergence of new antimicrobial-resistant bacteria and global dissemination of them and their antimicrobial resistance genes into animal and human populations. Efforts to prevent antimicrobial overuse in aquaculture must include education of all stakeholders about its detrimental effects on the health of fish, human beings, and the aquatic ecosystem (the notion of One Health), and encouragement of environmentally friendly measures of disease prevention, including vaccines, probiotics, and bacteriophages. Adoption of these measures is a crucial supplement to efforts dealing with antimicrobial resistance by developing new therapeutic agents, if headway is to be made against the increasing problem of antimicrobial resistance in human and veterinary medicine.

Borrelia chilensis, a new member of the Borrelia burgdorferi sensu lato complex that extends the range of this genospecies in the Southern Hemisphere

Borrelia burgdorferi sensu lato (s.l.), transmitted by Ixodes spp. ticks, is the causative agent of Lyme disease. Although Ixodes spp. ticks are distributed in both Northern and Southern Hemispheres, evidence for the presence of B. burgdorferi s.l. in South America apart from Uruguay is lacking. We now report the presence of culturable spirochetes with flat-wave morphology and borrelial DNA in endemic Ixodes stilesi ticks collected in Chile from environmental vegetation and long-tailed rice rats (Oligoryzomys longicaudatus). Cultured spirochetes and borrelial DNA in ticks were characterized by multilocus sequence typing and by sequencing five other loci (16S and 23S ribosomal genes, 5S-23S intergenic spacer, flaB, ospC). Phylogenetic analysis placed this spirochete as a new genospecies within the Lyme borreliosis group. Its plasmid profile determined by polymerase chain reaction and pulsed-field gel electrophoresis differed from that of B. burgdorferi B31A3. We propose naming this new South American member of the Lyme borreliosis group B. chilensis VA1 in honor of its country of origin.

Invasion and lysis of HeLa cell monolayers by Salmonella typhi: the role of lipopolysaccharide

Adhesion to and penetration of HeLa cell monolayers by Salmonella typhi Ty2 requires the presence of a complete lipopolysaccharide as demonstrated by the inability of polysaccharide-defective mutants to invade the monolayer. Lysis of HeLa cell monolayers by Salmonella typhi Ty2 is associated with intracellular bacterial multiplication and no detectable production of extracellular toxins. The ability of Salmonella typhi to invade and lyse monolayers could provide a novel system for the study of its ability to invade the bloodstream from the intestine.

The role of the stringent response in the pathogenesis of bacterial infections

Abstract Precise regulatory control of bacterial gene activation is crucial for the adaptation of pathogens to the hostile microniches of their hosts. The role of one of these global regulators, the stringent response, in pathogen virulence and persistence has recently become a favoured area of research. The complexity and multiplicity of the bacterial genes and regulatory pathways affected by the stringent response suggest that the relationship between the stringent response and virulence could be considerably more complex than expected and is perhaps unique for each pathogen and for each set of virulence properties that it regulates.