Ribonanza: deep learning of RNA structure through dual crowdsourcing.

Prediction of RNA structure from sequence remains an unsolved problem, and progress has been slowed by a paucity of experimental data. Here, we present Ribonanza, a dataset of chemical mapping measurements on two million diverse RNA sequences collected through Eterna and other crowdsourced initiatives. Ribonanza measurements enabled solicitation, training, and prospective evaluation of diverse deep neural networks through a Kaggle challenge, followed by distillation into a single, self-contained model called RibonanzaNet. When fine tuned on auxiliary datasets, RibonanzaNet achieves state-of-the-art performance in modeling experimental sequence dropout, RNA hydrolytic degradation, and RNA secondary structure, with implications for modeling RNA tertiary structure.

Frequency and Molecular Characterization of Staphylococcus aureus from Placenta of Mothers with Term and Preterm Deliveries.

Globally, prematurity is the leading cause of neonatal mortality (babies in the first four weeks of life) and now the second leading cause of mortality after pneumonia in children under age five. The neonatal gut microbial colonization is crucial in the human life cycle. Placental microbiota transmits from the gut microbiota plays a significant role in association with kinship. Simultaneously, this transition is being made from mother to infant. This comparative study explored the diversity of microbiota associated with term and preterm neonates by evaluating the placental samples. The study found that 16/68 (23.5%) full-term placental samples were positive for S. aureus; on the other hand, 4/16 (25%) preterm placental samples confirmed culture growth for S. aureus. Antimicrobial susceptibility patterns showed that Staphylococcusaureus (S. aureus) isolates from both types of samples were resistant to Ofloxacin, Trimethoprim-sulfamethoxazole, Oxacillin, and Cefoxitin. However, Methicillin-Resistant Staphylococcus aureus (MRSA) detection was 43.75% in full-term and 75% in preterm placental samples. Moreover, two isolates were positive for both mecA and PVL virulent genes, and the rest were positive only for the mecA gene. Interestingly few isolates lacked both characteristic MRSA genes, mecA and PVL. Notably, resistances were more inclined towards preterm samples for antimicrobial susceptibility and MRSA screening. It may be concluded that there is a significant presence of S. aureus in the placenta of mothers with term and preterm deliveries which might be responsible for preterm deliveries. Therefore, judicious use of antibiotics during pregnancies may help prevent preterm births.

Reviving Phage Therapy for the Treatment of Cholera.

Cholera remains a major risk in developing countries, particularly after natural or man-made disasters. Vibrio cholerae El Tor is the most important cause of these outbreaks, and is becoming increasingly resistant to antibiotics, so alternative therapies are urgently needed. In this study, a single bacteriophage, Phi_1, was used to control cholera prophylactically and therapeutically in an infant rabbit model. In both cases, phage-treated animals showed no clinical signs of disease, compared with 69% of untreated control animals. Bacterial counts in the intestines of phage-treated animals were reduced by up to 4 log10 colony-forming units/g. There was evidence of phage multiplication only in animals that received a V. cholerae challenge. No phage-resistant bacterial mutants were isolated from the animals, despite extensive searching. This is the first evidence that a single phage could be effective in the treatment of cholera, without detectable levels of resistance. Clinical trials in human patients should be considered.

High-Speed Atomic Force Microscopy Visualization of the Dynamics of the Multienzyme Fatty Acid Synthase.

Multienzymes, such as the protein metazoan fatty acid synthase (FAS), are giant and highly dynamic molecular machines for critical biosynthetic processes. The molecular architecture of FAS was elucidated by static high-resolution crystallographic analysis, while electron microscopy revealed large-scale conformational variability in FAS with some correlation to functional states in catalysis. However, little is known about time scales of conformational dynamics, the trajectory of motions in individual FAS molecules, and the extent of coupling between catalysis and structural changes. Here, we present an experimental single-molecule approach to film immobilized or selectively tethered FAS in solution at different viewing angles and high spatiotemporal resolution using high-speed atomic force microscopy. Mobility of individual regions of the multienzyme is recognized in video sequences, and correlation of shape features implies a convergence of temporal resolution and velocity of FAS dynamics. Conformational variety can be identified and grouped by reference-free 2D class averaging, enabling the tracking of conformational transitions in movies. The approach presented here is suited for comprehensive studies of the dynamics of FAS and other multienzymes in aqueous solution at the single-molecule level.

Bacteremia: Prevalence and antimicrobial resistance profiling in chronic kidney diseases and renal transplant patients.

OBJECTIVE: To determine the prevalence of bacteraemia in patients with chronic kidney disease, particularly those on dialysis and those who had had renal transplant, and to evaluate resistance among the isolated strains. METHODS: The cross-sectional study was conducted at Kidney Centre, Al-Sayyed Hospital, Rawalpindi, Pakistan, from June to December 2014. Samples that displayed positive growth were separated from the rest. The isolates were then identified and screened for extended spectrum beta lactamases and metallo beta lactamases production and other resistance mechanisms by phenotypic method. RESULTS: Of the 1400 samples, only 46 samples (3.3%) displayed signal for positive growth. The prevalence of extended spectrum beta lactamase (ESBL) producing strains was recorded to be 37%.Carbapenem resistance was witnessed in 15% samples. Whereas, Methicillin-resistant staphylococcus aureus prevalence was detected to be 2%. CONCLUSIONS: Resistance in gram-negative microbes was rising, while it was declining in gram-positive microbes.

Characterization and application of roxithromycin loaded cyclodextrin based nanoparticles for treatment of multidrug resistant bacteria.

An outbreak of infections with a high mortality rate caused by multidrug resistant (MDR) bacteria is one of the biggest health challenges globally. A class IV drug, roxithromycin (ROX), has poor solubility. In this study, ROX was first encapsulated in the cavity of each of the ß-cyclodextrin (ßCD) and hydroxypropyl-ß-cyclodextrin (HPßCD). Then, each of the resulting ßCD-ROX inclusion complex and HPßCD-ROX inclusion complex were separately loaded into poly-(lactic-co-glycolic acid) (PLGA) to synthesize ßCD-ROX/PLGA and HPßCD-ROX/PLGA nanoparticles (NPs). Blank and ROX loaded PLGA (ROX-PLGA) NPs were also prepared. The loading efficiency of ROX is comparatively high for HPßCD-ROX/PLGA NPs in comparison to the ßCD-ROX/PLGA NPs and ROX-PLGA NPs. All designed formulations showed significant (P<0.0001) antibacterial activity against the selected MDR bacterial strains. In a nutshell, this study demonstrated a great therapeutic potential of the above-mentioned delivery systems for treatment of MDR bacteria.

Mechanism of foreign DNA recognition by a CRISPR RNA-guided surveillance complex from Pseudomonas aeruginosa.

The Type I-F CRISPR-mediated (clustered regularly interspaced short palindromic repeats) adaptive immune system in Pseudomonas aeruginosa consists of two CRISPR loci and six CRISPR-associated (cas) genes. Foreign DNA surveillance is performed by a complex of Cas proteins (Csy1­4) that assemble with a CRISPR RNA (crRNA) into a 350-kDa ribonucleoprotein called the Csy complex. Here, we show that foreign nucleic acid recognition by the Csy complex proceeds through sequential steps, initiated by detection of two consecutive guanine­cytosine base pairs (G­C/G­C) located adjacent to the complementary DNA target. We show that this motif, called the PAM (protospacer adjacent motif), must be double-stranded and that single-stranded PAMs do not provide significant discriminating power. Binding assays performed with G­C/G­C-rich competitor sequences indicate that the Csy complex interacts directly with this dinucleotide motif, and kinetic analyses reveal that recognition of a G­C/G­C motif is a prerequisite for crRNA-guided binding to a target sequence. Together, these data indicate that the Csy complex first interacts with G­C/G­C base pairs and then samples adjacent target sequences for complementarity to the crRNA guide.

Evolutionary origins of the multienzyme architecture of giant fungal fatty acid synthase.

Fungal fatty acid synthase (fFAS) is a key paradigm for the evolution of complex multienzymes. Its 48 functional domains are embedded in a matrix of scaffolding elements, which comprises almost 50% of the total sequence and determines the emergent multienzymes properties of fFAS. Catalytic domains of fFAS are derived from monofunctional bacterial enzymes, but the evolutionary origin of the scaffolding elements remains enigmatic. Here, we identify two bacterial protein families of noncanonical fatty acid biosynthesis starter enzymes and trans-acting polyketide enoyl reductases (ERs) as potential ancestors of scaffolding regions in fFAS. The architectures of both protein families are revealed by representative crystal structures of the starter enzyme FabY and DfnA-ER. In both families, a striking structural conservation of insertions to scaffolding elements in fFAS is observed, despite marginal sequence identity. The combined phylogenetic and structural data provide insights into the evolutionary origins of the complex multienzyme architecture of fFAS.

Identification of possible virulence marker from Campylobacter jejuni isolates.

A novel protein translocation system, the type-6 secretion system (T6SS), may play a role in virulence of Campylobacter jejuni. We investigated 181 C. jejuni isolates from humans, chickens, and environmental sources in Vietnam, Thailand, Pakistan, and the United Kingdom for T6SS. The marker was most prevalent in human and chicken isolates from Vietnam.