- Volume 6, Issue 10, 2024

We report the draft genome assembly of strain 4866-2_S43 isolated from a eucalyptus lesion in Argentina and what until recently was caused by Xanthomonas citri pv. eucalyptorum (Xce). The genome size is 5 188 607 bp, with a G+C content of 64.66%. Comparative analysis reveals that the closest relative of strain 4866-2_S43 is Xce LPF 602, isolated in Brazil. Comparison of the whole genome sequences revealed an average nucleotide identity (ANI) of 99.96% between the two strains. ANIs were determined between the whole genome sequence of strain 4866-2_S43 and the genomes of all currently validated Xanthomonas spp. These results revealed that strain 4866-2_S43 shared >95% similarity with X. citri pv. citri and X. citri pv. phaseoli, and <95% with X. euvesicatoria pv. alfalfae, X. euvesicatoria pv. perforans, and X. euvesicatoria pathovars euvesicatoria and eucalyptii.

During the 3 years following the emergence of the COVID-19 pandemic, the African continent, like other regions of the world, was substantially impacted by COVID-19. In Morocco, the COVID-19 pandemic has been marked by the emergence and spread of several SARS-CoV-2 variants, leading to a substantial increase in the incidence of infections and deaths. Nevertheless, the comprehensive understanding of the genetic diversity, evolution, and epidemiology of several viral lineages remained limited in Morocco. This study sought to deepen the understanding of the genomic epidemiology of SARS-CoV-2 through a retrospective analysis. The main objective of this study was to analyse the genetic diversity of SARS-CoV-2 and identify distinct lineages, as well as assess their evolution during the pandemic in Morocco, using genomic epidemiology approaches. Furthermore, several key mutations in the functional proteins across different viral lineages were highlighted along with an analysis of the genetic relationships amongst these strains to better understand their evolutionary pathways. A total of 2274 genomic sequences of SARS-CoV-2 isolated in Morocco during the period of 2020 to 2023, were extracted from the GISAID EpiCoV database and subjected to analysis. Lineages and clades were classified according to the nomenclature of GISAID, Nextstrain, and Pangolin. The study was conducted and reported in accordance with STROBE (Strengthening the Reporting of Observational Studies in Epidemiology) guidelines. An exhaustive analysis of 2274 genomic sequences led to the identification of 157 PANGO lineages, including notable lineages such as B.1, B.1.1, B.1.528, and B.1.177, as well as variants such as B.1.1.7, B.1.621, B.1.525, B.1.351, B.1.617.1, B.1.617.2, and its notable sublineages AY.33, AY.72, AY.112, AY.121 that evolved over time before being supplanted by Omicron in December 2021. Among the 2274 sequences analysed, Omicron and its subvariants had a prevalence of 59.5%. The most predominant clades were 21K, 21L, and 22B, which are respectively related phylogenetically to BA.1, BA.2, and BA.5. In June 2022, Morocco rapidly observed a recrudescence of cases of infection, with the emergence and concurrent coexistence of subvariants from clade 22B such as BA.5.2.20, BA.5, BA.5.1, BA.5.2.1, and BF.5, supplanting the subvariants BA.1 (clade display 21K) and BA.2 (clade display 21L), which became marginal. However, XBB (clade 22F) and its progeny such XBB.1.5(23A), XBB.1.16(23B), CH.1.1(23C), XBB.1.9(23D), XBB.2.3(23E), EG.5.1(23F), and XBB.1.5.70(23G) have evolved sporadically. Furthermore, several notable mutations, such as H69del/V70del, G142D, K417N, T478K, E484K, E484A, L452R, F486P, N501Y, Q613H, D614G, and P681H/R, have been identified. Some of these SARS-CoV-2 mutations are known to be involved in increasing transmissibility, virulence, and antibody escape. This study has identified several distinct lineages and mutations involved in the genetic diversity of Moroccan isolates, as well as the analysis of their evolutionary trends. These findings provide a robust basis for better understanding the distinct mutations and their roles in the variation of transmissibility, pathogenicity, and antigenicity (immune evasion/reinfection). Furthermore, the noteworthy number of distinct lineages identified in Morocco highlights the importance of maintaining continuous surveillance of COVID-19. Moreover, expanding vaccination coverage would also help protect patients against more severe clinical disease.

Clostridioides difficile is the leading cause of antibiotic-associated infections worldwide. Within the host, C. difficile can transition from a sessile to a motile state by secreting PPEP-1, which releases the cells from the intestinal epithelium by cleaving adhesion proteins. PPEP-1 belongs to the group of Pro-Pro endopeptidases (PPEPs), which are characterized by their unique ability to cleave proline–proline bonds. Interestingly, another putative member of this group, CD1597, is present in C. difficile. Although it possesses a domain similar to other PPEPs, CD1597 displays several distinct features that suggest a markedly different role for this protein. We investigated the proteolytic activity of CD1597 by testing various potential substrates. In addition, we investigated the effect of the absence of CD1597 by generating an insertional mutant of the cd1597 gene. Using the cd1597 mutant, we sought to identify phenotypic changes through a series of in vitro experiments and quantitative proteomic analyses. Furthermore, we aimed to study the localization of this protein using a fluorogenic fusion protein. Despite its similarities to PPEP-1, CD1597 did not show proteolytic activity. In addition, the absence of CD1597 caused an increase in various sporulation proteins during the stationary phase, yet we did not observe any alterations in the sporulation frequency of the cd1597 mutant. Furthermore, a promoter activity assay indicated a very low expression level of cd1597 in vegetative cells, which was independent of the culture medium and growth stage. The low expression was corroborated by our comprehensive proteomic analysis of the whole cell cultures, which failed to identify CD1597. However, an analysis of purified C. difficile spores identified CD1597 as part of the spore proteome. Hence, we predict that the protein is involved in sporulation, although we were unable to define a precise role for CD1597 in C. difficile.

Abstract

The objective is to analyse the physicochemical properties, spatial structure and protein–protein interactions (PPIs) of L7/L12 protein using bioinformatics methods and predict their B- and T-cell epitopes to lay a theoretical foundation for developing a novel multiepitope vaccine (MEV). The National Center for Biotechnology Information (NCBI) database was searched for the amino acid sequences of L7/L12 from Brucella melitensis. In addition, the online softwares, ProtParam and ProtScale, were used to predict the physicochemical properties: NetPhos3.1 and CD-search to predict the phosphorylation sites and conserved domains; SOMPA and SWISS-MODEL to predict the secondary and tertiary structures; the STRING database to analyse the PPIs; and the IEDB, ABCpred, SVMTrip and SYFPEITHI databases to predict the B- and T-cell epitopes. L7/L12 was docked to Toll-like receptor 4 (TLR4), B-cell receptor (BCR), Major histocompatibility complex I-T cell receptor (MHC I-TCR) and MHC II-TCR complexes, respectively, and the binding ability of L7/L12 to the targeted receptors was tested. L7/L12, consisting of 124 amino acids, was determined to be a stable, intracellular, hydrophilic protein containing 6 phosphorylation sites and ribosomal protein-related conserved domains. α-helices accounted for 70.16 %, β-turns for 2.42 %, extended strands for 8.87 % and irregular coils for 18.55 % of the secondary structure. The PPIs indicated that L7/L12 was involved in the constitution of ribosomes and regulating the accuracy of the translation process. Three B-cells, two cytotoxic T lymphocytes and three helper T lymphocyte epitopes were finally screened by comparing multiple databases. L7/L12 binds to TLR4, BCR, MHC I-TCR and MHC II-TCR complexes and forms stable hydrogen bonds, respectively. L7/L12, which governs the translation curate of proteins, possesses several potentially advantageous epitopes, laying a theoretical foundation for designing MEVs.

Background. Among the most significant yet often ignored health issues worldwide are trauma and accidental injuries. India accounts for 11% of global deaths in road accidents, the highest in the world, according to the World Bank report. There are limited data about the bacterial contamination of road traffic accident (RTA) wounds and their antibiotic susceptibility patterns.

Materials and Methods. This prospective study was conducted in a tertiary care centre in northern India from January 2023 to January 2024. Wound deep swabs or aspirates were collected from RTA patients with traumatic injuries at different time intervals. Gram stain and culture were performed, and positive aerobic culture was subjected to antibiotic susceptibility testing. Organism identification was done using MALDI-TOF MS and routine biochemical tests. Blood samples were also collected to rule out bloodstream infections during follow-up if the patient became febrile or showed symptoms of systemic infection. Sepsis was defined in those patients who had two or more scores in the systemic inflammatory response syndrome criteria with a positive microbiological culture. Risk factors were evaluated for sepsis on the basis of the patient’s vitals, injury characteristics, procalcitonin, Glasgow Coma Scale (GCS) score, need for mechanical ventilation and complete blood count, which were obtained from the patient’s admission file.

Results. A total of 189 wound samples were collected, of which 99 (52.38%) samples showed the growth of microorganisms. The aerobic isolates included 69 (69.69%) Gram-negative bacilli, of which the majority were Klebsiella pneumoniae, 28 (28.28%) Gram-positive cocci, of which the majority were Staphylococcus aureus and 2 (2.02%) anaerobic isolates. Among the Gram-negative isolates, none of the isolates were resistant to colistin. All S. aureus isolates were susceptible to vancomycin, teicoplanin and levonadifloxacin. Sepsis developed in 50 (26.45 %) patients. Significant risk factors evaluated for sepsis were a raised procalcitonin level, a low GCS score, a higher injury severity score, the need for mechanical ventilation and a raised quick sequential organ failure assessment score.

Conclusion. It is essential to ascertain the profile of microorganisms isolated from RTA wounds in order to reduce antibiotic resistance and deliver efficient treatment.

Fluoroquinolone-resistant Escherichia coli sequence type (ST)1193 is a profound, emerging lineage associated with systemic, urinary tract and neonatal infections. Humans, companion animals and the environment are reservoirs for ST1193, which has been disseminated globally. Following its detection in 2007, ST1193 has been identified repeatedly amongst fluoroquinolone-resistant clones in Australia. However, despite the growing importance of ST1193, only three complete genomes are published in the literature, none of which are from Australia. Here we expand on the available ST1193 resources with the complete genomes of five ST1193 strains sequenced using Oxford Nanopore Technologies and Illumina. Using in silico genotyping, we found that all strains were multi-drug resistant, including resistances to fluoroquinolones and cephalosporins. In vitro antibiotic susceptibility testing mostly correlated with individual genotypes. The exception was MS8320, which had additional in vitro resistance to piperacillin/tazobactam, ampicillin/sulbactam, cefazolin and doripenem (carbapenem). Further investigation identified seven additional copies of an IS26 transposable unit carrying a bla TEM-1B beta-lactamase gene, suggesting this tandem amplification is associated with extended resistance phenotypes. Uropathogenicity factors, including three separate siderophore-encoding loci, were conserved in chromosomal and plasmid regions. Using all complete genomes, we further elucidated the recombination events surrounding the previously described K5/K1 capsular locus switch. Phenotypic confirmation of differing capsules in Australian ST1193 strains, coupled with genetic analysis revealing insertions downstream of the capsular locus, underscored the genetic distinctions between K5 and K1 capsule encoding strains. This study provides five new reference ST1193 genomes from Australia. These include the earliest complete K5-capsule ST1193 genomes on record (collected 2007), alongside our reference genome (MS10858), a clinical isolate obtained early during the ST1193 expansion and representative of the predominant K1-associated clade. These findings lay the foundations for further genomic and molecular analyses that may help understand the underlying reasons for the rapid global expansion of ST1193.

High-copy-number plasmids are indispensable tools for gene overexpression studies in prokaryotes to engineer pathways or probe phenotypes of interest. The development of genetic tools for the industrially relevant Actinobacteria is of special interest, given their utility in producing keratolytic enzymes and biologically active natural products. Within the Actinobacteria, Streptomyces–Escherichia coli shuttle vectors based on the SCP2* and pIJ101 incompatibility groups are widely employed for molecular cloning and gene expression studies. Here, the sequences of two commonly used pIJ101-based Streptomyces–E. coli shuttle vectors, pEM4 and pUWL201, were determined using next-generation sequencing. These plasmids drive the expression of heterologous genes using the constitutive ermE*p promoter. pEM4 was found to be 8.3 kbp long, containing a β-lactamase gene, thiostrepton resistance marker, the lacZɑ fragment, a ColE1 origin of replication and the Streptomyces pIJ101 origin of replication. pUWL201 was found to be 6.78 kbp long, containing a β-lactamase gene, thiostrepton resistance marker, the lacZɑ fragment, a ColE1 origin of replication and the Streptomyces pIJ101 origin of replication. Interestingly, the sequences for both pEM4 and pUWL201 exceed their previously reported size by 1.1 and 0.4 kbp, respectively. This report updates the literature with the corrected sequences for these shuttle vectors, ensuring their compatibility with modern synthetic biology cloning methodologies.

Introduction. Peritonitis is characterized by acute inflammation of the peritoneum, often resulting from digestive organ perforation or intra-abdominal septic focus. It may be of either infectious or non-infectious origin. The bacteria involved are those of the digestive flora (Enterobacteriaceae and anaerobes), while Gram-positive cocci and yeasts can be isolated in nosocomial infections. Our study aims to isolate and identify the germs involved in community-acquired peritonitis in order to assess their susceptibility to the antibiotics available in our country.

Methods. This is a retrospective study of the bacteriological profile of community peritonitis in Rabat Morocco. A total of 150 adult patients with peritonitis were admitted and samples were collected intraoperatively for bacteriological examination between 1 July 2022 and 30 April 2023.

Results. Among the 150 patients, 101 (67.8%) were males and 49 (32.2%) were females, with a sex ratio (M/F) of 2 : 1. The mean age of the patients was 40.5 years±20.12. The distribution of bacteria was dominated by Escherichia coli (44%). Overall, 70% of E. coli isolated exhibited resistance to ampicillin, whereas no resistance to ampicillin has been reported for Enterococcus.

Discussion. In the present study, we were interested in the bacteriological profile of community peritonitis in order to adapt the antibiotic therapy to our bacterial ecology. Our findings indicate a concerning trend of increasing resistance among E. coli to the commonly used amoxicillin/clavulanic acid combination in our clinical setting.

Conclusion. Consequently, there is a need to reassess the empiric antibiotic prescribed for the management of community-acquired peritonitis.

Introduction. Bloodstream infection (BSI) is a common complication with a high fatality rate in cancer patients. There are notable variations in the epidemiology of BSI over time and among different countries. Infections due to multidrug-resistant organisms (MDROs) such as extended-spectrum beta-lactamases (ESBLs) and carbapenem-resistant Enterobacteriaceae (CRE) are increasing. This may lead to inadequate empirical antibiotic therapy, increasing the antimicrobial resistance (AMR) problem and unfavourable outcomes in these immunocompromised patients. There is paucity of data pertaining to AMR in such vulnerable patients from developing countries such as India. The aim of this study was to investigate the distribution of the bacterial pathogens causing BSI and the AMR trend in cancer patients in central India.

Methodology. This single-centre retrospective observational study was conducted in a tertiary care cancer hospital. Patients with solid organ and haematological malignancies, both adults and paediatric, who had blood cultures sent to the microbiology laboratory from January 2018 to December 2022 were included. Blood cultures were processed using the BacT/ALERT 3D system (bioMérieux, France), and the identification of the bacteria and their antimicrobial susceptibility (AST) was performed using the Vitek 2 compact system (bioMérieux, France). Electronic medical records and microbiology lab records were used to retrieve the demographic and microbiological data. Microsoft Excel (RRID:SCR_016137) was used to enter and tabulate the data. Statistical analysis was performed using SPSS version 29 (RRID:SCR_002865).

Results. A total of 687 isolates from 524 patients were studied. Gram-negative bacteria (64%) were the commonest cause of BSI in the studied patients, followed by Gram-positive cocci (25%) and fungal isolates (9%). Ten cases were polymicrobial. Escherichia coli (n=140) was the most common among the isolated pathogens, followed by Klebsiella species (n=103), Pseudomonas species (n=102), and coagulase-negative staphylococci (CONS) (n=92). Among the 140 isolates of E. coli, 66% were extended-spectrum β-lactamase (ESBL) producers and 26% were resistant to carbapenem. Among the 103 isolated Klebsiella species, 50% were carbapenem resistant and 36% were ESBL producers. Among enterobacterales, the CRE rate was 34%. Carbapenem resistance was seen in 25% of Pseudomonas species and 53% of Acinetobacter species isolates. Klebsiella species were the most resistant pathogens isolated. CONS comprised 56% of all Gram-positive isolates, followed by Staphylococcus aureus (36%), enterococci species (11%), and streptococci species (3%). Methicillin resistance was 60% in CONS and 64% in S. aureus. One vancomycin-resistant enterococcus was isolated. Non-albicans Candida was the most common fungal pathogen. The sensitivity to fluconazole was 84% in non-albicans Candida species, while only one isolate of Candida albicans was resistant to fluconazole. The trend of pathogens was insignificant over 5 years, with Gram-negative bacteria being the commonest. Further, there was no significant change in the trend of ESBL and CRE resistance pattern over 5 years.

Conclusion. Gram-negative bacteria were the most common isolated pathogens from BSI with a higher antimicrobial resistance rate in cancer patients. The CRE rate of 34% is alarming, limiting the choices for empirical antibiotic therapy.

Lactiplantibacillus pentosus is a probiotic bacterium reported to be present in various fermented foods, such as fermented olives, and it significantly influences human health. The present study concerns a lactic acid bacterial strain designated L. pentosus krglsrbmofpi2, isolated from traditional fermented rice, and which has been shown to have an assortment of beneficial attributes. Using Illumina technologies, we have sequenced and investigated the whole genome sequence of L. pentosus krglsrbmofpi2 to understand its functionality and safety. The chromosomal genome was 3.7 Mb in size with 46% GC content and 3192 protein-coding genes. Additional extensive bioinformatics investigations were carried out involving whole genome sequence assembly and annotation.

Respiratory infections account for millions of hospital admissions worldwide. The aetiology of respiratory infections can be attributed to a diverse range of pathogens including viruses, bacteria and fungi. SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2)-negative specimens from Wattansoppeng city, South Sulawesi, were analysed to study the spectrum of respiratory viruses. Samples were screened for influenza virus, enterovirus, Paramyxoviridae, Nipah virus, Coronaviridae and Pneumoviridae. Of 210 specimens, 19 were positive for respiratory syncytial virus (RSV)-A, RSV-B, human parainfluenza virus type 1 (HPIV-1), HPIV-2, human rhinovirus (HRV)-A, HRV-B, HRV-C, human metapneumovirus (HMPV), influenza A virus (IAV) and coxsackievirus A6 (CV-A6). Influenza virus was of seasonal H3N2 subtype. The HMPVs were of genotypes B1 and A2a, while one RSV-A was of the ON-1 genotype. The viruses mostly affected children with unknown severity.

Multiple transboundary animal diseases (TADs) circulate in Plateau State, Nigeria, where livestock keeping is common and contributes to both the physical and socio-economic well-being of a large proportion of the population. In this study, we explored the potential for environmental sampling to detect viruses causing TADs circulating in the region. Electrostatic dust cloths were used to swab areas of the environment likely to have contact with secretions and excretions from infected animals. Samples were collected monthly from five households, one transhumance site and one livestock market in two local government areas in Plateau State between March and October 2021. These were tested for the presence of peste des petits ruminants virus (PPRV) and capripox viruses using real-time PCR. Of the 458 samples collected, 2.4% (n = 11) were positive for PPRV RNA and 1.3 % (n = 6) were positive for capripox virus DNA. A capripox differentiation assay showed that these samples were positive for sheep pox virus (n = 2), goat pox virus (n = 2) and lumpy skin disease virus (n = 2). Our results demonstrate that environmental sampling could be used as part of TAD surveillance in the area. Environmental swabs require little technical knowledge to collect and can be used to detect multiple viruses from a single sample.

We report a case of cutaneous melioidosis in a 54-year-old male with a meropenem-resistant sub-population. He was empirically treated with episodic doxycycline and trimethoprim–sulfamethoxazole; however, the abscess re-accumulated. The patient had no prior exposure to meropenem. A sub-population of the isolate was meropenem resistant with an MIC >32 µg ml−1 and the identification was re-confirmed as Burkholderia pseudomallei. Whole-genome sequencing with ARDaP analysis only revealed a resistance determinant to doxycycline and did not reveal a resistance determinant to meropenem. Furthermore, no carbapenemases were detected through multiple bioinformatics tools. To date, this is the first reported case in Australia of a B. pseudomallei isolate resistant to meropenem without previous carbapenem exposure.

Non-typhoidal Salmonella lung infections are rare and are usually confined to immunocompromised hosts. Previous case reports have found that usually patients have either gastroenteritis or bacteraemia in addition to pulmonary involvement. We present the first known reported case of a Salmonella Weltevreden lung abscess and empyema in an immunocompetent patient without gastroenteritis. Despite the use of antimicrobials active against the pathogen, the patient needed surgical intervention to achieve adequate source control. While S. Weltevreden has previously been associated with returned travellers, especially from Southeast Asia, its incidence in Queensland is now increasing. Therefore, it is important for clinicians to be aware of its potential severity as well as the range of presentations.

Peritoneal dialysis is a blood purification technique used in cases of end-stage chronic kidney failure, based on the filtering capabilities of the peritoneum. Infections, often caused by poor asepsis during catheter manipulation, are generally attributed to Staphylococcus epidermidis and Staphylococcus aureus. Corynebacterium, usually considered non-pathogenic, is rarely involved in these infections. We present a case of peritonitis due to Corynebacterium amycolatum in a patient undergoing peritoneal dialysis. The diagnosis was made based on cytobacteriological examination of the dialysate fluid, which on two occasions showed high levels of white blood cells with a predominance of neutrophilic polymorphonuclear and a monomorphic appearance of colonies on agar medium, whose identification by biochemical tests and antibiotic sensitivity study confirmed the presence of C. amycolatum. The patient was successfully treated with vancomycin, resulting in symptom resolution and sterilization of the dialysate fluid. Although rare, the involvement of Corynebacterium species underscores the importance of confirming its pathogenicity. Further studies are needed to better understand the epidemiology of these infections and guide future treatments. This case also highlights the need for a rigorous approach to confirming the pathogenicity of Corynebacterium despite its traditional classification as a contaminant.