The Role of Substrate Mediated Allostery in the Catalytic Competency of the Bacterial Oligosaccharyltransferase PglB

The oligosaccharyltransferase of Campylobacter lari (PglB) catalyzes the glycosylation of asparagine in the consensus sequence N-X-S/T, where X is any residue except proline. Molecular dynamics simulations of PglB bound to two different substrates were used to characterize the differences in the structure and dynamics of the substrate-enzyme complexes that can explain the higher catalytic efficiency observed for substrates containing threonine at the +2 position rather than serine. We observed that a threonine-containing substrate is more tightly bound than a serine-containing substrate. Because serine lacks a methyl group relative to threonine, the serine-containing peptide cannot stably form simultaneous van der Waals interactions with T316 and I572 as the threonine-containing substrate can. As a result, the peptide-PglB interaction is destabilized and the allosteric communication between the periplasmic domain and external loop EL5 is disrupted. These changes ultimately lead to the reorientation of the periplasmic domain relative to the transmembrane domain such that the two domains are further apart compared to PglB bound to the threonine-containing peptide. The crystal structure of PglB bound to the peptide and a lipid-linked oligosaccharide analog shows a pronounced closing of the periplasmic domain over the transmembrane domain in comparison to structures of PglB with peptide only, indicating that a closed conformation of the domains is needed for catalysis. The results of our studies suggest that lower enzymatic activity observed for serine versus threonine results from a combination of less stable binding and structural changes in PglB that influence the ability to form a catalytically competent state. This study illustrates a mechanism for substrate specificity via modulation of dynamic allosteric pathways.

Prevalence and Antibiotic Resistance of Campylobacter spp. in Urban and Rural Black-Headed Gulls Chroicocephalus ridibundus

We investigate the role of black-headed gulls (Chroicocephalus ridibundus), an omnivorous species that is among the most likely wild bird candidates for transmission of zoonotic agents, as a potential reservoir of Campylobacter spp. Colonies with different anthropogenic pressures were studied to examine differences in exposure to sources of Campylobacter between rural and urban birds. We recorded Campylobacter spp. in 4.87% of adult black-headed gulls and 2.22% of their chicks after analysing 1036 cloacal swabs collected over two breeding seasons in three colonies in northern Poland. Campylobacter jejuni was found most frequently (85.72%), and Campylobacter lari and Campylobacter coli were much scarcer. Prevalence of Campylobacter did not differ significantly between black-headed gulls breeding in urban (4.27%) and rural (3.80%) habitats. Almost all isolates from chicks and adults were susceptible to azithromycin (97.62%) and erythromycin (95.24%), but fewer to tetracycline (50.00%) and ciprofloxacin (47.62%). Campylobacter prevalence was unrelated to the date of sampling. Our study indicates that black-headed gulls are carriers of resistant to antibiotics Campylobacter and they can contaminate natural waterbodies with their faeces, which poses a threat to human and farm animal health.

Multidrug-resistant Campylobacter jejuni, Campylobacter coli and Campylobacter lari isolated from asymptomatic school-going children in Kibera slum, Kenya

Background: The objective of this study was to determine the prevalence of thermophilic Campylobacter spp. in asymptomatic school-going children and establish the antibiotic resistance patterns of the isolates towards the drugs used to treat campylobacteriosis, including macrolides, quinolones and tetracycline. Campylobacter spp. are a leading cause of enteric illness and have only recently shown resistance to antibiotics. Methods: This study isolated Campylobacter spp., including Campylobacter coli, Campylobacter jejuni and Campylobacter lari, in stool samples from asymptomatic school-going children in one of the biggest urban slums in Kenya. The disc diffusion method using EUCAST breakpoints was used to identify antibiotic-resistant isolates, which were further tested for genes encoding for tetracycline resistance using primer-specific polymerase chain reaction. Results: In total, 580 stool samples were collected from 11 primary schools considering both gender and age. Subjecting 294 biochemically characterized Campylobacter spp. isolates to genus-specific PCR, 106 (18.27% of stool samples) isolates were confirmed Campylobacter spp. Out of the 106 isolates, 28 (4.83%) were Campylobacter coli, 44 (7.58%) were Campylobacter jejuni while 11 (1.89%) were Campylobacter lari. Campylobacter jejuni had the highest number of isolates that were multi-drug resistant, with 26 out of the 28 tested isolates being resistant to ciprofloxacin (5 mg), nalidixic acid (30 mg), tetracycline (30 mg) and erythromycin (15 mg). Conclusions: In conclusion, asymptomatic school going children in the study area were found to be carriers of multidrug resistant Campylobacter coli, Campylobacter jejuni and Campylobacter lari at 84%. A one-health approach, which considers overlaps in environment, animals and human ecosystems, is recommended in addressing multidrug resistane in Campylobacter, since animals are the main reservoirs and environmental contamination is evident.

Multidrug-resistant Campylobacter jejuni, Campylobacter coli and Campylobacter lari isolated from asymptomatic school-going children in Kibera slum, Kenya

Background: The objective of this study was to determine the prevalence of thermophilic Campylobacter spp. in asymptomatic school-going children and establish the antibiotic resistant patterns of the isolates towards the drugs used to treat campylobacteriosis, including macrolides, quinolones and tetracycline. Campylobacter spp. are a leading cause of enteric illness and have only recently shown resistant to antibiotics. Methods: This study isolated Campylobacter spp., including Campylobacter coli, Campylobacter jejuni and Campylobacter lari, in stool samples from asymptomatic school-going children in one of the biggest urban slums in Kenya. The disc diffusion method using EUCAST breakpoints was used to identify antibiotic-resistant isolates, which were further tested for genes encoding for tetracycline resistances using primer-specific polymerase chain reaction. Results: In total, 580 stool samples were collected from 11 primary schools considering both gender and age. Subjecting 294 biochemically characterized Campylobacter spp. isolates to genus-specific PCR, 106 (18.27% of stool samples) isolates were confirmed Campylobacter spp. Out of the 106 isolates, 28 (4.83%) were Campylobacter coli, 44 (7.58%) were Campylobacter jejuni while 11 (1.89%) were Campylobacter lari. Campylobacter jejuni had the highest number of isolates that were multi-drug resistant, with 26 out of the 28 tested isolates being resistant to ciprofloxacin (5 mg), nalidixic acid (30 mg), tetracycline (30 mg) and erythromycin (15 mg). Conclusions: In conclusion, a one-health approach, which considers overlaps in environment, animals and human ecosystems, is recommended in addressing campylobacteriosis in humans, since animals are the main reservoirs and environmental contamination is evident.