Laparoscopic Cholecystectomy with a Mixed Approach in a Patient with Kartagener Syndrome: Technical Report and Review of Literature

Kartagener syndrome (KS) is a rare autosomal recessive disease. The disease is characterized by three typical symptoms: chronic sinusitis, situs viscerum inversus (SVI), and bronchiectasis. The laparoscopic cholecystectomy (LC) is the standard procedure in most cases of cholelithiasis, but in SVI patients it can be difficult, especially for right-handed surgeons. We report the case of a 24-year-old female affected by KS, presenting with a history of symptomatic cholelithiasis. Ultrasound and magnetic resonance cholangiopancreatography confirmed SVI totalis and cholelithiasis. The patient underwent a laparoscopic cholecystectomy by a right-handed surgeon performed with a mixed approach without complications. Laparoscopic cholecystectomy in SVI patients can be a safe and reliable technique especially for a left-handed surgeon. The described technique is also easy for a right-handed surgeon. However, it is considered a technically challenging procedure and often requires technical modification

Multiplex PCR to detect bacteriophages from natural whey cultures of buffalo milk and characterisation of two phages active against Lactococcus lactis, фApr-1 and фApr-2

This work investigated bacteriophage induced starter failures in artisanal buffalo Mozzarella production plants in Southern Italy. Two hundred and ten samples of whey starter cultures were screened for bacteriophage infection. Multiplex polymerase chain reaction (PCR) revealed phage infection in 28.56% of samples, all showing acidification problems during cheese making. Based on DNA sequences, bacteriophages for Lactococcus lactis (L. lactis), Lactobacillus delbruekii (L. delbruekii) and Streptococcus thermophilus (S. thermophilus) were detected. Two phages active against L.  lactis, фApr-1 and фApr-2, were isolated and characterised. The genomes, approximately 31.4 kb and 31 kb for фApr-1 and фApr-2 respectively, consisted of double-stranded linear DNA with pac-type system. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) showed one major structural protein of approximately 32.5 kDa and several minor proteins. This is the first report of phage isolation in buffalo milk and of the use of multiplex PCR to screen and study the diversity of phages against Lactic Acid Bacteria (LAB) strains in artisanal Water Buffalo Mozzarella starters

Campylobacter bacteriophage DA10: An excised temperate bacteriophage targeted by CRISPR-cas

Background: Lytic bacteriophages that infect Campylobacter spp. have been utilized to develop therapeutic/decontamination techniques. However, the association of Campylobacter spp. and bacteriophages has been the focus of several strands of research aimed at understanding the complex relationships that have developed between predators and prey over evolutionary time. The activities of endogenous temperate bacteriophages have been used to evaluate genomic rearrangements and differential protein expression in host cells, and mechanisms of resistance to bacteriophage infection in campylobacters such as phase variation and CRISPR-mediated immunity. Results: Temperate bacteriophage DA10 represents a novel excised and infective virus capable of replication in a restricted set of C. jejuni and C. coli hosts. Whole genome sequencing reveals that DA10 (35,379 bp) forms part of a novel group of temperate bacteriophages that have limited distribution among database host genome sequences. Analysis of potential host genomes reveals a robust response against DA10 and DA10-like bacteriophages is driven by CRISPR-mediated immunity with 75% of DA10 ORFs represented as ~ 30 bp spacer sequences in numerous Campylobacter Type II-C CRISPR arrays. Several DA10-like homologues have been identified in a small sub-set of C. jejuni and C. coli genome sequences (ranging from near complete integrated prophage sequences to fragments recognisable in the sequence read archive). Conclusions: A complete intact DA10-like prophage in C. jejuni CJ677CC520 provides evidence that the associations between host and DA10-like bacteriophages are long-standing in evolutionary timescales. Extensive nucleotide substitution and loss can be observed in the integrated DA10-like prophage of CJ677CC520 compared to other relatives as observed through pairwise genome comparisons. Examining factors that have limited the population expansion of the prophage, while others appear to have thrived and prospered (Mu-like, CJIE-like, and lytic Campylobacter bacteriophages) will assist in identifying the underlying evolutionary processes in the natural environment

Single Primer Enrichment Technology (SPET) for High-Throughput Genotyping in Tomato and Eggplant Germplasm

[EN] Single primer enrichment technology (SPET) is a new, robust, and customizable solution for targeted genotyping. Unlike genotyping by sequencing (GBS), and like DNA chips, SPET is a targeted genotyping technology, relying on the sequencing of a region flanking a primer. Its reliance on single primers, rather than on primer pairs, greatly simplifies panel design, and allows higher levels of multiplexing than PCR-based genotyping. Thanks to the sequencing of the regions surrounding the target SNP, SPET allows the discovery of thousands of closely linked, novel SNPs. In order to assess the potential of SPET for high-throughput genotyping in plants, a panel comprising 5k target SNPs, designed both on coding regions and introns/UTRs, was developed for tomato and eggplant. Genotyping of two panels composed of 400 tomato and 422 eggplant accessions, comprising both domesticated material and wild relatives, generated a total of 12,002 and 30,731 high confidence SNPs, respectively, which comprised both target and novel SNPs in an approximate ratio of 1:1.6, and 1:5.5 in tomato and eggplant, respectively. The vast majority of the markers was transferrable to related species that diverged up to 3.4 million years ago (Solanum pennellii for tomato and S. macrocarpon for eggplant). Maximum Likelihood phylogenetic trees and PCA outputs obtained from the whole dataset highlighted genetic relationships among accessions and species which were congruent with what was previously reported in literature. Better discrimination among domesticated accessions was achieved by using the target SNPs, while better discrimination among wild species was achieved using the whole SNP dataset. Our results reveal that SPET genotyping is a robust, high-throughput technology for genetic fingerprinting, with a high degree of cross-transferability between crops and their cultivated and wild relatives, and allows identification of duplicates and mislabeled accessions in genebanks.This work has been funded by the European Union's Horizon 2020 Research and Innovation Programme under the grant agreement number 677379 (G2P-SOL project: Linking genetic resources, genomes, and phenotypes of solanaceous crops).Barchi, L.; Acquadro, A.; Alonso-Martín, D.; Aprea, G.; Bassolino, L.; Demurtas, O.; Ferrante, P.... (2019). Single Primer Enrichment Technology (SPET) for High-Throughput Genotyping in Tomato and Eggplant Germplasm. Frontiers in Plant Science. 10:1-17. https://doi.org/10.3389/fpls.2019.01005S11710Acquadro, A., Barchi, L., Gramazio, P., Portis, E., Vilanova, S., Comino, C., … Lanteri, S. (2017). 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Cross-sectional study of hepatitis E virus (HEV) circulation in Italian pig farms

OHEJP Project: BIOPIGEE Foodborne transmission is considered the main way of spreading zoonotic hepatitis E virus (HEV) infection in Europe. In recent years, the human cases of hepatitis E in subjects without history of travel in endemic areas have raised, suggesting that domestic HEV transmission is increasing. Pork products with or without liver, are often indicated as the source of many human foodborne HEV cases as well as small outbreaks. Pigs are recognized as the main reservoir of the zoonotic HEV-3 genotype, the most frequently detected in human cases in the EU. In the absence of a harmonized surveillance of HEV circulation, data on prevalence are heterogeneous but confirm a widespread circulation of HEV-3 in pig herds across EU. HEV-3 can pass through the food chain from farm to fork when infected animals are slaughtered. In Italy, several studies reported the circulation of HEV-3 in pig farms, but results are heterogeneous due to dierent methodologies applied. In the present study, we performed a survey over 51 pig herds belonging to three main types of farms: breeding, fattening and farrow-to- finish. HEV-RNA was analyzed by broad range Real-time RT-PCR on 20 samples for each farm, obtained by pooling together feces from 10 individuals. Overall, HEV RNA was confirmed on 150 fecal pooled samples out of 1,032 (14.5%). At least one positive pooled sample was detected from 18 farms out of 51 tested (35.3%). By lowering the number of infected pigs at primary production, the risk of HEV-3 entering into the food chain can be reduced. Hence, information on HEV circulation in herds is highly relevant for choosing preventive measures and deserves development of a monitoring program and further investigations

Bacteriophage therapy to reduce colonization of campylobacter jejuni in broiler chickens before slaughter

Campylobacteriosis is the most commonly reported gastrointestinal disease in humans. Campybacter jejuni is the main cause of the infection, and bacterial colonization in broiler chickens is widespread and difficult to prevent, leading to high risk of occurrence in broiler meat. Phage therapy represents an alternative strategy to control Campylobacter in poultry. The aim of this work was to assess the efficacy of two field-isolated bacteriophages against experimental infections with an anti-microbial resistant (AMR) Campylobacter jejuni strain. A two-step phage application was tested according to a specific combination between chickens’ rearing time and specific multiplicities of infections (MOIs), in order to reduce the Campylobacter load in the animals at slaughtering and to limit the development of phage-resistant mutants. In particular, 75 broilers were divided into three groups (A, B and C), and phages were administered to animals of groups B and C at day 38 (Φ 16-izsam) and 39 (Φ 7-izsam) at MOI 0.1 (group B) and 1 (group C). All broilers were euthanized at day 40, and Campylobacter jejuni was enumerated in cecal contents. Reductions in Campylobacter counts were statistically significant in both group B (1 log10 colony forming units (cfu)/gram (gr)) and group C (2 log10 cfu/gr), compared to the control group. Our findings provide evidence about the ability of phage therapy to reduce the Campylobacter load in poultry before slaughtering, also associated with anti-microbial resistance pattern

The rapid spread of SARS-COV-2 Omicron variant in Italy reflected early through wastewater surveillance

The SARS-CoV-2 Omicron variant emerged in South Africa in November 2021, and has later been identified worldwide, raising serious concerns. A real-time RT-PCR assay was designed for the rapid screening of the Omicron variant, targeting characteristic mutations of the spike gene. The assay was used to test 737 sewage samples collected throughout Italy (19/21 Regions) between 11 November and 25 December 2021, with the aim of assessing the spread of the Omicron variant in the country. Positive samples were also tested with a real-time RT-PCR developed by the European Commission, Joint Research Centre (JRC), and through nested RT-PCR followed by Sanger sequencing. Overall, 115 samples tested positive for Omicron SARS-CoV-2 variant. The first occurrence was detected on 7 December, in Veneto, North Italy. Later on, the variant spread extremely fast in three weeks, with prevalence of positive wastewater samples rising from 1.0% (1/104 samples) in the week 5–11 December, to 17.5% (25/143 samples) in the week 12–18, to 65.9% (89/135 samples) in the week 19–25, in line with the increase in cases of infection with the Omicron variant observed during December in Italy. Similarly, the number of Regions/Autonomous Provinces in which the variant was detected increased fromone in the first week, to 11 in the second, and to 17 in the last one. The presence of the Omicron variant was confirmed by the JRC real-time RT-PCR in 79.1% (91/115) of the positive samples, and by Sanger sequencing in 66% (64/97) of PCR amplicons