Phage Milagro: a platform for engineering a broad host range virulent phage for Burkholderia.

IMPORTANCE: Burkholderia infections are a significant concern in people with CF and other immunocompromising disorders, and are difficult to treat with conventional antibiotics due to their inherent drug resistance. Bacteriophages, or bacterial viruses, are now seen as a potential alternative therapy for these infections, but most of the naturally occurring phages are temperate and have narrow host ranges, which limit their utility as therapeutics. Here we describe the temperate Burkholderia phage Milagro and our efforts to engineer this phage into a potential therapeutic by expanding the phage host range and selecting for phage mutants that are strictly virulent. This approach may be used to generate new therapeutic agents for treating intractable infections in CF patients.

An Experimental Field Trial Investigating the Use of Bacteriophage and Manure Slurry Applications in Beef Cattle Feedlot Pens for Salmonella Mitigation.

Post-harvest Salmonella mitigation techniques are insufficient at addressing Salmonella harbored in cattle lymph nodes, necessitating the exploration of pre-harvest alternatives that reduce Salmonella prior to dissemination to the lymph nodes. A 2 × 2, unbalanced experiment was conducted to determine the effectiveness of pre-harvest treatments applied to the pen surface for Salmonella mitigation in cattle. Treatments included manure slurry intended to mimic pen run-off water (n = 4 pens), a bacteriophage cocktail (n = 4), a combination of both treatments (n = 5), and a control group (n = 5) that received no treatment. Environment samples from 18 feedlot pens and fecal grabs, hide swabs, and subiliac lymph nodes from 178 cattle were collected and selectively enriched for Salmonella, and Salmonella isolates were sequenced. The combination treatment was most effective at reducing Salmonella, and the prevalence was significantly lower compared with the control group for rump swabs on Days 14 and 21. The treatment impact on Salmonella in the lymph nodes could not be determined due to low prevalence. The reduction on cattle hides suggests that bacteriophage or water treatments applied to the feedlot pen surface may reduce Salmonella populations in cattle during the pre-harvest period, resulting in reduced contamination during slaughter and processing.

Compounding Achromobacter Phages for Therapeutic Applications.

Achromobacter species colonization of Cystic Fibrosis respiratory airways is an increasing concern. Two adult patients with Cystic Fibrosis colonized by Achromobacter xylosoxidans CF418 or Achromobacter ruhlandii CF116 experienced fatal exacerbations. Achromobacter spp. are naturally resistant to several antibiotics. Therefore, phages could be valuable as therapeutics for the control of Achromobacter. In this study, thirteen lytic phages were isolated and characterized at the morphological and genomic levels for potential future use in phage therapy. They are presented here as the Achromobacter Kumeyaay phage collection. Six distinct Achromobacter phage genome clusters were identified based on a comprehensive phylogenetic analysis of the Kumeyaay collection as well as the publicly available Achromobacter phages. The infectivity of all phages in the Kumeyaay collection was tested in 23 Achromobacter clinical isolates; 78% of these isolates were lysed by at least one phage. A cryptic prophage was induced in Achromobacter xylosoxidans CF418 when infected with some of the lytic phages. This prophage genome was characterized and is presented as Achromobacter phage CF418-P1. Prophage induction during lytic phage preparation for therapy interventions require further exploration. Large-scale production of phages and removal of endotoxins using an octanol-based procedure resulted in a phage concentrate of 1 × 109 plaque-forming units per milliliter with an endotoxin concentration of 65 endotoxin units per milliliter, which is below the Food and Drugs Administration recommended maximum threshold for human administration. This study provides a comprehensive framework for the isolation, bioinformatic characterization, and safe production of phages to kill Achromobacter spp. in order to potentially manage Cystic Fibrosis (CF) pulmonary infections.

Assessment of availability, readiness, and challenges for scaling-up hypertension management services at primary healthcare facilities, Central Highland region, Vietnam, 2020.

INTRODUCTION: Vietnam aims for 95% of commune health stations (CHSs) to have functional hypertension management programs by 2025. However, limited resources may impede the Central Highland region health system from achieving this goal. We assessed the availability and readiness of hypertension management services at CHSs in the Central Highland region and identified challenges to facilitate evidence-based planning. METHODS: We used a mixed-methods cross-sectional design to assess hypertension management services using WHO's service availability and readiness assessment (SARA) tools in all 579 CHSs in the region, combined with twenty in-depth interviews of hypertension program focal points at communal, district, and provincial levels in all four provinces. We descriptively analyzed quantitative data and thematically analyzed qualitative data. RESULTS: Hypertension management services were available at 65% of CHSs, and the readiness of the services was 62%. The urban areas had higher availability and readiness indices in most domains (basic amenities, basic equipment, and essential medicines) compared to rural areas, except for staff and training. The qualitative results showed a lack of trained staff and ambiguity in national hypertension treatment guidelines, insufficient essential medicines supply mechanism, and low priority and funding limitations for the hypertension program. CONCLUSION: The overall availability and readiness for hypertension diagnosis and management service at CHSs in the Central Highland region were low, reflecting inadequate capacity of the primary healthcare facilities. Some measures to strengthen hypertension programs in the region might include increased financial support, ensuring a sufficient supply of basic medicines, and providing more specific treatment guidelines.

Competitive interference among rhizobia reduces benefits to hosts.

The capacity of beneficial microbes to compete for host infection-and the ability of hosts to discriminate among them-introduces evolutionary conflict that is predicted to destabilize mutualism. We investigated fitness outcomes in associations between legumes and their symbiotic rhizobia to characterize fitness impacts of microbial competition. Diverse Bradyrhizobium strains varying in their capacity to fix nitrogen symbiotically with a common host plant, Acmispon strigosus, were tested in full-factorial coinoculation experiments involving 28 pairwise strain combinations. We analyzed the effects of interstrain competition and host discrimination on symbiotic-interaction outcomes by relativizing fitness proxies to clonally infected and uninfected controls. More than one thousand root nodules of coinoculated plants were genotyped to quantify strain occupancy, and the Bradyrhizobium strain genome sequences were analyzed to uncover the genetic bases of interstrain competition outcomes. Strikingly, interstrain competition favored a fast-growing, minimally beneficial rhizobia strain. Host benefits were significantly diminished in coinoculation treatments relative to expectations from clonally inoculated controls, consistent with competitive interference among rhizobia that reduced both nodulation and plant growth. Competition traits appear polygenic, linked with inter-strain allelopathic interactions in the rhizosphere. This study confirms that competition among strains can destabilize mutualism by favoring microbes that are superior in colonizing host tissues but provide minimal benefits to host plants. Moreover, our findings help resolve the paradox that despite efficient host control post infection, legumes nonetheless encounter rhizobia that vary in their nitrogen fixation.

Therapeutic Potential of Intravenous Phage as Standalone Therapy for Recurrent Drug-Resistant Urinary Tract Infections.

Recurrent urinary tract infections (rUTI) are common in kidney transplant recipients (KTR) and are associated with multidrug resistance and increased morbidity/mortality. Novel antibiotic alternatives to reduce UTI recurrence are critically needed. We describe a case of rUTI due to extended spectrum beta lactamase (ESBL) Klebsiella pneumoniae in a KTR that was treated successfully with 4 weeks of adjunctive intravenous bacteriophage therapy alone, without concomitant antibiotics, and with no recurrence in a year of follow-up.

Yap and Taz promote osteogenesis and prevent chondrogenesis in neural crest cells in vitro and in vivo.

Neural crest cells (NCCs) are multipotent stem cells that can differentiate into multiple cell types, including the osteoblasts and chondrocytes, and constitute most of the craniofacial skeleton. Here, we show through in vitro and in vivo studies that the transcriptional regulators Yap and Taz have redundant functions as key determinants of the specification and differentiation of NCCs into osteoblasts or chondrocytes. Primary and cultured NCCs deficient in Yap and Taz switched from osteogenesis to chondrogenesis, and NCC-specific deficiency for Yap and Taz resulted in bone loss and ectopic cartilage in mice. Yap bound to the regulatory elements of key genes that govern osteogenesis and chondrogenesis in NCCs and directly regulated the expression of these genes, some of which also contained binding sites for the TCF/LEF transcription factors that interact with the Wnt effector ß-catenin. During differentiation of NCCs in vitro and NCC-derived osteogenesis in vivo, Yap and Taz promoted the expression of osteogenic genes such as Runx2 and Sp7 but repressed the expression of chondrogenic genes such as Sox9 and Col2a1. Furthermore, Yap and Taz interacted with ß-catenin in NCCs to coordinately promote osteoblast differentiation and repress chondrogenesis. Together, our data indicate that Yap and Taz promote osteogenesis in NCCs and prevent chondrogenesis, partly through interactions with the Wnt-ß-catenin pathway.

Comparative genomics of Acinetobacter baumannii and therapeutic bacteriophages from a patient undergoing phage therapy.

In 2016, a 68-year-old patient with a disseminated multidrug-resistant Acinetobacter baumannii infection was successfully treated using lytic bacteriophages. Here we report the genomes of the nine phages used for treatment and three strains of A. baumannii isolated prior to and during treatment. The phages used in the initial treatment are related, T4-like myophages. Analysis of 19 A. baumannii isolates collected before and during phage treatment shows that resistance to the T4-like phages appeared two days following the start of treatment. We generate complete genomic sequences for three A. baumannii strains (TP1, TP2 and TP3) collected before and during treatment, supporting a clonal relationship. Furthermore, we use strain TP1 to select for increased resistance to five of the phages in vitro, and identify mutations that are also found in phage-insensitive isolates TP2 and TP3 (which evolved in vivo during phage treatment). These results support that in vitro investigations can produce results that are relevant to the in vivo environment.

Complete Genome Sequence of Stenotrophomonas maltophilia Phage Philippe.

Stenotrophomonas maltophilia is emerging as an opportunistic multidrug-resistant pathogen. S. maltophilia podophage Philippe has a 74,717-bp genome which is related broadly to the N4-like phage group, including Stenotrophomonas phage Pokken. The low sequence identity to other described phages suggests that Philippe is an unclassified member of the N4-like subfamily Rothmandenesvirinae.

Complete Genome Sequence of Stenotrophomonas maltophilia Siphophage Siara.

Stenotrophomonas maltophilia is associated with an increasing incidence of nosocomial infections. Here, we describe the isolation and genome annotation of S. maltophilia siphophage Siara. Its 61,427-bp genome is currently related only to one phage in the NCBI database, namely, S. maltophilia phage Salva, and is not related to any prophages.

Complete Genome Sequence of Stenotrophomonas maltophilia Podophage Pepon.

Stenotrophomonas maltophilia is an opportunistic bacterium that is commonly associated with respiratory infections in immunocompromised patients, including cystic fibrosis patients. In this report, we introduce the complete genome sequence of S. maltophilia podophage Pepon, which is a T7-like phage closely related to the previously reported phage Ponderosa.

Complete Genome Sequence of Stenotrophomonas maltophilia Podophage Paxi.

Stenotrophomonas maltophilia is a multidrug-resistant nosocomial pathogen that can cause life-threatening infections among immunocompromised populations. This report presents the complete 74,962-bp genome of S. maltophilia podophage Paxi, an N4-like phage sharing 85.3% nucleotide similarity to S. maltophilia podophage Pokken.

Complete Genome Sequence of Stenotrophomonas maltophilia Siphophage Suso.

Phage Suso is a temperate siphophage of Stenotrophomonas maltophilia with a 44,659-bp genome. This phage is closely related to Stenotrophomonas phage SM171, sharing 92% overall nucleotide identity as determined by BLASTn, and it shares 14 similar proteins (BLASTp, E value < 0.001) with some Pseudomonas phages from the genus Beetrevirus.

Complete Genome Sequence of Stenotrophomonas maltophilia Siphophage Sonora.

Phage Sonora is a siphophage that was isolated against the opportunistic human pathogen Stenotrophomonas maltophilia. The genome of phage Sonora is 63,825 bp long and is not related to that of any phage at the nucleotide level. Sonora shares 46 of 97 total proteins with the Bordetella phages CN2, MW2, and FP1.

Complete Genome Sequence of Stenotrophomonas maltophilia Podophage Piffle.

Stenotrophomonas maltophilia is an emerging multidrug-resistant opportunistic human pathogen causing various nosocomial infections. Here, we characterize the genome of S. maltophilia podophage Piffle. Its 76,332-bp genome is most closely related to the N4-like S. maltophilia podophage Pokken, with over 86% genome-wide nucleotide identity and 84 shared proteins.

Complete Genome Sequence of Burkholderia cenocepacia Phage Paku.

Burkholderia cenocepacia is able to cause infections in cystic fibrosis patients. B. cenocepacia phage Paku has a 42,727-bp genome sharing a phiKMV-like genome arrangement. T7-like tail components were identified in parallel with a tyrosine integrase, suggesting that Paku might exhibit a temperate lifestyle, an atypical feature for an Autographiviridae phage.

Complete Genome Sequence of Enterococcus faecalis Siphophage Sigurd.

Enterococcus faecalis is associated with antibiotic-resistant infections, and this study presents E. faecalis siphophage Sigurd. The 41,811-bp Sigurd genome is divided into two arms defined by long convergent predicted transcription units that are separated by a bidirectional rho-independent terminator. Sigurd has a small terminase that is closely related to Bacillus subtilis cos phage phi105.

Complete Genome Sequence of Alcaligenes faecalis Phage Piluca.

Alcaligenes faecalis is an opportunistic pathogen exhibiting drug resistance. Here, the 35,451-bp genome of A. faecalis phage Piluca is described. Piluca is not closely related to any isolated phages in the NCBI database. Piluca possesses genes encoding CI-like and Cro-like repressors and a tyrosine integrase, suggesting its temperate lifestyle.

Complete Genome Sequence of Stenotrophomonas maltophilia Podophage Ptah.

Stenotrophomonas maltophilia is an opportunistic pathogen demonstrating increasing drug resistance. Here, the genome of the T7-like S. maltophilia podophage Ptah is described. Its 42,593-bp genome is closely related to previously reported T7-like S. maltophilia podophages, including phage Ponderosa.