Bacteriophage ZCKP1: a potential treatment for Klebsiella pneumoniae isolated from diabetic foot patients

The recorded growth in infection by multidrug resistant bacteria necessitates prompt efforts towards developing alternatives to antibiotics, such as bacteriophage therapy. Immuno-compromised patients with diabetes mellitus are particularly prone to foot infections by multidrug resistant Klebsiella pneumoniae, which may be compounded by chronic osteomyelitis. Bacteriophage ZCKP1, isolated from freshwater in Giza, Egypt, was tested in vitro to evaluate its lytic activity against a multidrug resistant K. pneumoniae KP/01, isolated from foot wound of a diabetic patient in Egypt. Characterization of ZCKP1 phage indicated that it belonged to the Myoviridae family of bacteriophages with a ds-DNA genome size of 150.9 kb. Bacteriophage ZCKP1 lysed a range of osteomyelitis pathogenic agents including Klebsiella spp., Proteus spp. and E. coli isolates. The bacteriophage reduced the bacterial counts of host bacteria by ≥2 log10 CFU/ml at 25°C, and demonstrated the ability to reduce bacterial counts and biofilm biomass (> 50%) when applied at high multiplicity of infection (50 PFU/CFU). These characteristics make ZCKP1 phage of potential therapeutic value to treat K. pneumoniae and associated bacteria present in diabetic foot patients

High-throughput sequencing reveals genetic determinants associated with antibiotic resistance in Campylobacter spp. from farm-to-fork

[EN]Campylobacter species are one of the most common causative agents of gastroenteritis worldwide. Resistance against quinolone and macrolide antimicrobials, the most commonly used therapeutic options, poses a serious risk for campylobacteriosis treatment. Owing to whole genome sequencing advancements for rapid detection of antimicrobial resistance mechanisms, phenotypic and genotypic resistance trends along the “farm-to-fork” continuum can be determined. Here, we examined the resistance trends in 111 Campylobacter isolates (90 C. jejuni and 21 C. coli) recovered from clinical samples, commercial broiler carcasses and dairy products in Cairo, Egypt. Multidrug resistance (MDR) was observed in 10% of the isolates, mostly from C. coli. The prevalence of MDR was the highest in isolates collected from broiler carcasses (13.3%), followed by clinical isolates (10.5%), and finally isolates from dairy products (4%). The highest proportion of antimicrobial resistance in both species was against quinolones (ciprofloxacin and/or nalidixic acid) (68.4%), followed by tetracycline (51.3%), then erythromycin (12.6%) and aminoglycosides (streptomycin and/or gentamicin) (5.4%). Similar resistance rates were observed for quinolones, tetracycline, and erythromycin among isolates recovered from broiler carcasses and clinical samples highlighting the contribution of food of animal sources to human illness. Significant associations between phenotypic resistance and putative gene mutations was observed, with a high prevalence of the gyrA T86I substitution among quinolone resistant isolates, tet(O), tet (W), and tet(32) among tetracycline resistant isolates, and 23S rRNA A2075G and A2074T mutations among erythromycin resistant isolates. Emergence of resistance was attributed to the dissemination of resistance genes among various lineages, with the dominance of distinctive clones. For example, sub-lineages of CC828 in C. coli and CC21 in C. jejuni and the genetically related clonal complexes ‘CC206 and CC48’ and ‘CC464, CC353, CC354, CC574’, respectively, propagated across different niches sharing semi-homogenous resistance patterns.SIThis work was partially funded by the Zewail City internal research fund (agreement number ZC 004-2019) and joint ASRT-BA research grant (project number 1110) awarded to Dr. Mohamed Elhadidy. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Characterization and comprehensive genome analysis of novel bacteriophage, vB_Kpn_ZCKp20p, with lytic and anti-biofilm potential against clinical multidrug-resistant Klebsiella pneumoniae

IntroductionThe rise of infections by antibiotic-resistant bacterial pathogens is alarming. Among these, Klebsiella pneumoniae is a leading cause of death by hospital-acquired infections, and its multidrug-resistant strains are flagged as a global threat to human health, which necessitates finding novel antibiotics or alternative therapies. One promising therapeutic alternative is the use of virulent bacteriophages, which specifically target bacteria and coevolve with them to overcome potential resistance. Here, we aimed to discover specific bacteriophages with therapeutic potential against multiresistant K. pneumoniae clinical isolates.Methods and ResultsOut of six bacteriophages that we isolated from urban and medical sewage, phage vB_Kpn_ZCKp20p had the broadest host range and was thus characterized in detail. Transmission electron microscopy suggests vB_Kpn_ZCKp20p to be a tailed phage of the siphoviral morphotype. In vitro evaluation indicated a high lytic efficiency (30 min latent period and burst size of ∼100 PFU/cell), and extended stability at temperatures up to 70°C and a wide range of (2-12) pH. Additionally, phage vB_Kpn_ZCKp20p possesses antibiofilm activity that was evaluated by the crystal violet assay and was not cytotoxic to human skin fibroblasts. The whole genome was sequenced and annotated, uncovering one tRNA gene and 33 genes encoding proteins with assigned functions out of 85 predicted genes. Furthermore, comparative genomics and phylogenetic analysis suggest that vB_Kpn_ZCKp20p most likely represents a new species, but belongs to the same genus as Klebsiella phages ZCKP8 and 6691. Comprehensive genomic and bioinformatics analyses substantiate the safety of the phage and its strictly lytic lifestyle.ConclusionPhage vB_Kpn_ZCKp20p is a novel phage with potential to be used against biofilm-forming K. pneumoniae and could be a promising source for antibacterial and antibiofilm products, which will be individually studied experimentally in future studies

The effect of the timing of exposure to Campylobacter jejuni on the gut microbiome and inflammatory responses of broiler chickens

Background Campylobacters are an unwelcome member of the poultry gut microbiota in terms of food safety. The objective of this study was to compare the microbiota, inflammatory responses, and zootechnical parameters of broiler chickens not exposed to Campylobacter jejuni with those exposed either early at 6 days old or at the age commercial broiler chicken flocks are frequently observed to become colonized at 20 days old. Results Birds infected with Campylobacter at 20 days became cecal colonized within 2 days of exposure, whereas birds infected at 6 days of age did not show complete colonization of the sample cohort until 9 days post-infection. All birds sampled thereafter were colonized until the end of the study at 35 days (mean 6.1 log10 CFU per g of cecal contents). The cecal microbiota of birds infected with Campylobacter were significantly different to age-matched non-infected controls at 2 days post-infection but generally the composition of the cecal microbiota were more affected by bird age as the time post infection increased. The effects of Campylobacter colonization on the cecal microbiota were associated with reductions in the relative abundance of OTUs within the taxonomic family Lactobacillaceae and the Clostridium cluster XIVa. Specific members of the Lachnospiraceae and Ruminococcaceae families exhibit transient shifts in microbial community populations dependent upon the age at which the birds become colonized by C. jejuni. Analysis of ileal and cecal chemokine/cytokine gene expression revealed increases in IL-6, IL-17A and Il-17F consistent with a Th17 response but the persistence of the response was dependent on the stage/time of C. jejuni colonization that coincide with significant reductions in the abundance of Clostridium cluster XIVa. Conclusions This study combines microbiome data, cytokine/chemokine gene expression with intestinal villus and crypt measurements to compare chickens colonized early or late in the rearing cycle to provide insights into the process and outcomes of Campylobacter colonization. Early colonization results in a transient growth rate reduction and pro-inflammatory response but persistent modification of the cecal microbiota. Late colonization produces pro-inflammatory responses with changes in the cecal microbiota that will endure in market ready chickens

Integral quality control studies of Campylobacter on poultry and poultry meat : occurrence, frequency and survival of Campylobacter and bacteriophage from free-range and organic broiler chickens

EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Enumeration and Diversity of Campylobacters and Bacteriophages Isolated during the Rearing Cycles of Free-Range and Organic Chickens

Campylobacters and Campylobacter-specific bacteriophages were isolated and enumerated during the rearing cycle of free-range (56 days) and organic chickens (73 days) at 3-day intervals from hatching until slaughter. In both flocks Campylobacter jejuni was the initial colonizer but Campylobacter coli was detected more frequently from 5 weeks of age. The diversity of the Campylobacter isolates was examined by pulsed-field gel electrophoresis of SmaI-digested genomic DNA and antimicrobial resistance typing. Bacteriophages were isolated from 51% (19 of 37 birds) of Campylobacter-positive organic birds (log(10) 2.5 to log(10) 5.7 PFU/g of cecal contents). The bacteriophages were all typical group III Campylobacter bacteriophages in terms of genomic size but could be characterized in terms of their host range and placed into five different groups. In contrast to the organic birds, anti-Campylobacter activity (bacteriocin-like) was observed in 26% (10 of 38 birds) of Campylobacter-positive free-range birds, and only one bacteriophage was isolated. Appearance of either bacteriophages or anti-Campylobacter activity was associated with changes in the levels of colonization and the predominant genotypes and species isolated. The frequency and potential influence of naturally occurring bacteriophages and/or inhibitory substances on the diversity and fluctuations of populations of campylobacters have not previously been reported in either free-range or organic chickens

Manufacture of Mudaffara Cheese. II. From Standardised Buffaloes Milk Fortified with Different Levels of Skim Milk Powder

Page(s): 2 (2), 92- 100, 9 Ref.Mudaffara cheese was made from standardised buffaloes' milk and buffaloes' milk fortified with 2.5% and 5% skim milk powder. A standard procedure for the production of Mudaffara cheese has been outlined. On addition of skim milk powder, the total solids of milk increased and consequently yield of cheese produced also increased. The chemical analysis of the product of the three milk samples showed significant differences in very few parameters, namely tyrosine and tryptophan contents of cheese. Organoleptic evaluation revealed no differences in all quality parameters tested for the three cheese samples (P<0.05). Skim milk powder could be added to buffaloes' milk at a rate of up to 5%, so as to increase yield, without affecting the chemical composition of the cheese or its sensory qualities.Khartoum University Pres

Using millimeter‐waves for rapid detection of pathogenic bacteria in food based on bacteriophage

The accessibility of a rapid method for detection and identification of food‐borne pathogens is crucial for food industry worldwide. Antibiotic resistance bacteria (eg, E. coli ) that can enter the food chain in different ways, can indeed survive on foods causing disease to humans. Hence, the introduction of a rapid detection technology becomes necessary for the food industry to ensure consumer safety, especially for products with short shelf lives. Bacteriophages can be used to detect and identify bacteria. In this study, a novel biosensor is proposed to detect pathogens by means of phage‐based baroreceptor. The biosensing technique is based on millimeter‐waves technology in the 30 to 60 GHz frequency range. The proposed biosensor can detect the pathogenic bacteria in different food samples by using a diamond‐shape microstrip slot antenna. The bacteriophage‐bacterium interaction is detected through the dynamic changes in transmission lines and antennas responses. The correctness of the antenna to detect E. coli in real food sample (tomato) is also investigated. The results indicate that, through the designed sensing elements, the transient interaction between bacteria and phage can effectively be detected. This sensing mechanism allows for a faster, more accurate, and low‐cost detection of pathogenic bacteria than traditional assays. Finally, the results are compared with previously reported sensing techniques