The integrase protein of the Rhizobium meliloti 41 phage 16-3 has been classified as a member of the Int family of tyrosine recombinases. The site-specific recombination system of the phage belongs to the group in which the target site of integration (attB) is within a tRNA gene. Since tRNA genes are conserved, we expected that the target sequence of the site-specific recombination system of the 16-3 phage could occur in other species and integration could take place if the required putative host factors were also provided by the targeted cells. Here we report that a plasmid (pSEM167) carrying the attP element and the integrase gene (int) of the phage can integrate into the chromosomes of R. meliloti 1021 and eight other species. In all cases integration occurred at so-far-unidentified, putative proline tRNA (CGG) genes, indicating the possibility of their common origin. Multiple alignment of the sequences suggested that the location of the att core was different from that expected previously. The minimal attB was identified as a 23-bp sequence corresponding to the anticodon arm of the tRNA.Site-specific integration is an efficient way to introduce new genetic information into the chromosome of the cell in a targeted fashion and ensure stable maintenance of the feature gained. Integrative vectors developed by utilizing site-specific integration elements from temperate bacteriophages (i.e., attP and the integrase gene combined) provide tools for this technology. Since the presence of the appropriate target sequence (attB) in the recipient chromosome and specific host factors are required for the integrative process, utilization of a particular integrative recombination system is frequently restricted to the host range of a given phage. A number of known eubacterial temperate phages, such as L5 (37), HP1 (24), 16-3 (35), RP3 (21), U (50), P4 (38), Mx8 (28), Sfi21 (7), 10MC (22), T12 (31), A2 (2), mv4 (16), Ms6 (19), and VWB (51), use tRNA genes as target sequences for integration into the host chromosomes. There are some plasmids with integrative functions, such as pSAM2 (30), pSE101 (5), pSE211 (6), and pSLP1 (52), whose target sites are also within tRNA genes. The SSV1 virus of the thermophilic archaeon Sulfolobus shibatae can also integrate into a tRNA (40, 41). Considering the conservative nature of tRNAs, one can expect that the target sites for a particular integrative system could occur in several bacterial species at various levels of relatedness. This view was confirmed by well-documented examples: the site-specific recombination systems of Ms6, mv4, VWB, pSAM2, pSE101, and pSLP1 were shown to function in closely related bacteria (3,5,19,39,51,52). Moreover A2 provides an example of integration in both gram-negative and gram-positive species (2).The temperate bacteriophage 16-3 of Rhizobium meliloti 41 has been studied thoroughly. In addition to genetic and physical characterization of its genome (10,12,14), the central region of the phage chromosome (GenBank accession no. AJ131679) was investigated in de...

The aim was to compare the prevalence of acute infection and seropositivity of SARS-CoV-2 among healthcare workers (HCWs) and medical students. Material and Methods: A high-volume, single-center analysis was conducted in the period of July 1-August 1, 2020, at the Semmelweis University. Naso-and oropharyngeal samples were collected for polymerase chain reaction (PCR), and blood samples for anti-SARS-CoV-2 IgG. A questionnaire was also administered about the infection symptoms and the obtained results were assessed by profession and site of care delivery. Results: From the total cohort (N = 7948), 4478 (56%) and 3470 (44%) were health professionals and medical students, respectively. They were mainly female (67%), and the mean age of HCWs and students was 40 and 25 years, respectively. By profession, physicians (1.5%) and other HCWs (1.8%) showed a comparable SARS-CoV-2 exposure. International students had the highest (2.1%), whereas Hungarian students had the lowest (0.6%) prevalence of seropositivity. The highest prevalence was detected among the staff of COVID-19 wards (12.1%). By PCR, medical students showed the lowest occurrence of active infection with a prevalence of 0.17%, while physicians and other HCWs had a higher prevalence (1.46% and 1.71%, respectively). By site of care delivery, positive test results were the most frequent at COVID-19 wards (3.8%). Conclusions: Physicians and other HCWs showed comparable SARS-CoV-2 seropositivity prevalence, approximately twice as high as in the general population of Budapest. Hungarian students had lower prevalence of seropositivity than this reference. High prevalence among international students suggests that they had imported the infection. The very high prevalence of documented exposure among staff members at COVID-19 wards urges for improving the safety measures.

SummarySeveral temperate bacteriophage utilize chromosomal sequences encoding putative tRNA genes for phage attachment. However, whether these sequences belong to genes which are functional as tRNA is generally not known. In this article, we demonstrate that the attachment site of temperate phage 16-3 ( attB ) nests within an active proline tRNA gene in Rhizobium meliloti 41. A loss-of-function mutation in this tRNA gene leads to significant delay in switching from lag to exponential growth phase. We converted the putative Rhizobium gene to an active amber suppressor gene which suppressed amber mutant alleles of genes of 16-3 phage and of Escherichia coli origin in R. meliloti 41 and in Agrobacterium tumefaciens GV2260. Upon lysogenization of R. meliloti by phage 16-3 , the proline tRNA gene retained its structural and functional integrity. Aspects of the co-evolution of a temperate phage and its bacterium host is discussed. The side product of this work, i.e. construction of amber suppressor tRNA genes in Rhizobium and Agrobacterium , for the first time widens the options of genetic study.

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