Biology subjects

Nissan, I.

Publications and source records attributed to Nissan, I..

6 recordsLinked to original sources

bioRxiv2026

HiSpoT: a novel one-tube dual-barcode PCR method for NGS spoligotyping amplicon library preparation in Mycobacterium tuberculosis complex

Tuberculosis remains one of the leading infectious causes of death globally, making rapid, accurate, and affordable strain typing essential for effective outbreak tracking and epidemiological surveillance. To address the limitations and high costs of current typing methods, we developed a novel one-tube, dual-barcode PCR method for preparing targeted next-generation sequencing (NGS) amplicon libraries for spoligotyping of Mycobacterium tuberculosis complex (MTBC), which we termed HiSpoT (High-throughput Spoligotyping by Targeted next generation sequencing). The assay targets 43 conserved spacer oligonucleotides in the CRISPR direct repeat (DR) region and is presented here together with the previously unpublished, difficult-to-design dual-barcode primer set needed to multiplex 96 samples in a single tube. HiSpoT is accurate, fast, high-throughput, and low-cost, eliminating the need for a commercial NGS library preparation kit. Because HiSpoT is PCR-based, the resulting spoligotypes remain directly comparable with the large PCR-based spoligotype databases accumulated over decades, enabling rapid outbreak tracking and biogeographic characterization of the tuberculosis burden of a country, while higher-resolution whole-genome sequencing (WGS) data are obtained. Analysis of 288 clinical MTBC isolates from Israel (2020-2022) demonstrated a broad range of genetic lineages, and comparison of 79 isolates analyzed by both HiSpoT and WGS showed 70% full concordance, with essentially all discrepancies attributable to single-spacer insertion-element disruptions of PCR primer sites. We also present two open-source Google Colab Python tools, Spoligotyping_MTBC_gcl and its condensed HiSpoT_Spoligotyping_v8_ready VIBE variant, freely available on GitHub, which automatically process Illumina paired-end fastq output from HiSpoT or WGS and report binary and octal spoligotypes, spoligo international types (SIT), and MTBC family assignments with accompanying quality-control reports.

molecular biology↗

bioRxiv2024

Enterobactin and Salmochelin S4 inhibit the growth of Staphylococcus aureus

There is increasing demand for novel antimicrobial agents to tackle the antimicrobial resistance crisis. Here we report that two Enterobacteriaceae-produced siderophores, enterobactin and salmochelin S4, inhibit the growth of Staphylococcus aureus isolates, including methicillin-resistance S. aureus (MRSA) clinical isolates. The MIC50 for different S. aureus isolates were 2-5 M for salmochelin S4 and 5-10 M for enterobactin. This inhibitory activity was partially repressed by adding Fe+3. These siderophores also inhibited the growth of Enterococcus strains, including vancomycin-resistant enterococci (VRE) clinical isolates, though less effectively than for S. aureus. The growth of various Gram-negative bacteria was barely affected by these siderophores. These results shed new light on the role of enterobactin and salmochelin in bacterial physiology and ecology and have potential for the development of novel strategies to combat the rapid rise of multidrug-resistant bacteria.

microbiology↗

bioRxiv2023

Legionella felix sp. - A novel Legionella species isolated in Israel from hot tap water

Bacteria of the genus Legionella are natural pathogens of the environment that can cause legionellosis, which can result in severe human pneumonia called Legionnaires disease. Here we describe a novel Legionella species isolated from hot tap water. High quality complete genome was generated using a combination of Nanopore and Illumina sequencing approaches. Our novel Legionella isolate possesses a 3,913,681 bp chromosome, (GC content 38.78% Mol) and a single novel large incF conjugative plasmid of 160,500 bp (GC content 37.97%). Interestingly, the chromosome encodes for 14 secondary metabolites biosynthetic gene clusters, more than any known other Legionella sp. The novel plasmid encodes for multiple genes that confer resistance to heavy metals. Bioinformatics analysis including average nucleotide identity (ANI) and genome-based taxonomy computation revealed that the genome of the new isolate differs from any previously described Legionella species. The closest related species to our isolate is Legionella cherrii. The name proposed for the new specie is Legionella felix in honor of Dr. Arthur Felix (1887-1956), a pioneering microbiologist, and member of the Royal Society of UK, who established the National Public Health Laboratory in Tel Aviv.

microbiology↗

bioRxiv2023

Type VII secretion system and its effect on Group B Streptococcus virulence

GBS may cause a devasting disease in newborns. In early onset disease of the newborn the bacteria are acquired from the colonized mother during delivery. We characterized type VII secretion system (T7SS), exporting small proteins of the WXG100 superfamily, in group B Streptococci (GBS) isolates from pregnant colonized women and newborns with early onset disease (EOD) to understand better understand T7SS contribution to virulence in these different clinical scenarios. GBS isolates were obtained from colonized mother prior to delivery and from newborns with EOD. DNA was analyzed for T7SS genes. A mutant EOD strain (ST17) was created by knocking out the essC gene encoding a T7SS protein. Galleria mellonella larvae were used to compare virulence of colonizing, EOD, and mutant EOD isolates. 33 GBS genomes were tested, 17 EOD isolates and 16 colonizing isolates. The T7SS locus encoded 8 genes: essC, membrane-embedded proteins (essA; essB), modulators of T7SS activity (esaA; esaB; esaC) and effectors: [esxA (SAG1039); esxB (SAG1030). ST17 isolates encode two copies of the essC gene and esxA gene encoding putative effectors but were present only in 23.5% of isolates. In ST1 isolates three copies of esxA gene were identified, but in ST6 and ST19 isolates all T7SS genes were missing. EOD isolates demonstrated enhanced virulence in G. mellonella model compared to colonizing isolates. The 118659{Delta}essC strain was attenuated in its killing ability, and the larvae were more effective in eradicating 118659{Delta}essC infection. essC gene deletion was associated with reduced bacterial growth. We demonstrated that T7SS plays an essential role during infection and contributes to GBS pathogenicity. Author SummaryType VII secretion system (T7SS) is related to virulence in various bacteria but is not well characterized in Group B Streptococci (GBS). GBS may cause sepsis, meningitis, and death in newborns. The bacteria rarely cause disease in pregnant mothers. Newborns acquire GBS from the colonized mother during delivery. We studied the role of T7SS in GBS isolates obtained from newborns with GBS sepsis in the first week of life and in colonized pregnant mothers. By studying T7SS genes we discovered that the genetic structure of the T7SS differs between isolates causing severe disease and colonizing isolates. To study the virulence of different GBS isolates we injected them into larvae and monitored larvae survival. Isolates causing severe disease in the newborn caused a more severe disease in larvae compared to colonizing isolates. We then deleted T7SS genes in GBS isolates causing severe disease. The killing activity of GBS isolates without T7SS genes was attenuated. The larva responded to these bacteria similarly to the response found when injecting the larva with GBS isolates from colonized mothers. These results support our hypothesis that T7SS is important for causing severe infection in the newborn and that this system contributes to GBS pathogenicity.

microbiology↗

bioRxiv2021

Systematic monitoring of photosynthetically-derived oxidative signals using a 2-Cys peroxiredoxin-based redox probe

Transmission of reductive cues from the photosynthetic electron transport chain to redox-regulated proteins plays a crucial role in activating chloroplast metabolism. However, deciphering the role of their counterbalanced oxidative signals is challenging due to monitoring difficulties. Here, we demonstrate the light-depended redox modification of chloroplast-targeted 2-Cys peroxiredoxins and introduce peroxiredoxin-based biosensors to monitor photosynthetically-derived oxidative signals. By employing a set of genetically encoded biosensors, we show the induction of oxidative signals under habitual light intensities and their inverse relationship with NADPH levels, unraveling the combined activity of reducing and oxidizing signals in fine-tuning chloroplast metabolism. A faster increase in carbon assimilation rates during photosynthesis induction phase was measured in plants deficient in 2-Cys peroxiredoxins compared to wild-type, suggesting the involvement of oxidative signals in attenuating photosynthesis under variable light environments. We suggest that oxidative signals measured by peroxiredoxin-based biosensors reflect the limitation to photosynthesis imposed by the redox regulatory system. One-Sentence SummaryA genetically encoded biosensor unmasked the dominant role of photosynthetically-derived oxidative signals under habitual conditions.

plant biology↗

bioRxiv2020

Mutations in the tomato gibberellin receptors suppresses xylem proliferation and reduces water loss under water-deficit conditions

Low gibberellin (GA) activity in tomato (Solanum lycopersicum) inhibits leaf expansion and reduces stomatal conductance. These lead to lower transpiration and improve water status under transient drought conditions. Tomato has three GIBBERELLIN-INSENSITIVE DWARF1 (GID1) GA receptors with overlapping activities and high redundancy. We have tested whether mutation in a single GID1 reduces transpiration without affecting growth and productivity. CRISPR-Cas9 gid1 mutants were able to maintain higher leaf water content under water-deficit conditions. Moreover, while gid1a exhibited normal growth, it showed reduced whole plant transpiration and better recovery from dehydration. Mutation in GID1a inhibited xylem vessels proliferation that led to lower hydraulic conductance. In stronger GA mutants, we also found reduced xylem vessel expansion. These results suggest that low GA activity affects transpiration by multiple mechanisms; it reduces leaf area, promotes stomatal closure and reduces xylem proliferation and expansion and as a result, xylem hydraulic conductance. We further examined if gid1a perform better than the control M82 in the field. Under these conditions, the high redundancy of GID1s was lost and gid1a plants were semi-dwarf, but their productivity was not affected. Although gid1a did not perform better under drought conditions in the field, it exhibited higher harvest index. HighlightThe loss of the tomato gibberellin receptors GID1s reduced xylem proliferation and xylem hydraulic conductance. These contribute to the effect of low gibberellin activity on water loss under water-deficit condition.

plant biology↗