Molecular characterization of Pseudomonas aeruginosa from diabetic foot infections in Tunisia.

Background. Pseudomonas aeruginosa is an invasive organism that frequently causes severe tissue damage in diabetic foot ulcers.Gap statement. The characterisation of P. aeruginosa strains isolated from diabetic foot infections has not been carried out in Tunisia.Purpose. The aim was to determine the prevalence of P. aeruginosa isolated from patients with diabetic foot infections (DFIs) in Tunisia and to characterize their resistance, virulence and molecular typing.Methods. Patients with DFIs admitted to the diabetes department of the International Hospital Centre of Tunisia, from September 2019 to April 2021, were included in this prospective study. P. aeruginosa were obtained from the wound swabs, aspiration and soft tissue biopsies during routine clinical care and were confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Antimicrobial susceptibility testing, serotyping, integron and OprD characterization, virulence, biofilm production, pigment quantification, elastase activity and molecular typing were analysed in all recovered P. aeruginosa isolates by phenotypic tests, specific PCRs, sequencing, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing.Results. Sixteen P. aeruginosa isolates (16.3 %) were recovered from 98 samples of 78 diabetic patients and were classified into 6 serotypes (O:11 the most frequent), 11 different PFGE patterns and 10 sequence types (three of them new ones). The high-risk clone ST235 was found in two isolates. The highest resistance percentages were observed to netilmicin (69 %) and cefepime (43.8 %). Four multidrug-resistant (MDR) isolates (25 %) were detected, three of them being carbapenem-resistant. The ST235-MDR strain harboured the In51 class 1 integron (intI1 +aadA6+orfD+qacED1-sul1). According to the detection of 14 genes involved in virulence or quorum sensing, 5 virulotypes were observed, including 5 exoU-positive, 9 exoS-positive and 2 exoU/exoS-positive strains. The lasR gene was truncated by ISPpu21 insertion sequence in one isolate, and a deletion of 64 bp in the rhlR gene was detected in the ST235-MDR strain. Low biofilm, pyoverdine and elastase production were detected in all P. aeruginosa; however, the lasR-truncated strain showed a chronic infection phenotype characterized by loss of serotype-specific antigenicity, high production of phenazines and high biofilm formation.Conclusions. Our study demonstrated for the first time the prevalence and the molecular characterization of P. aeruginosa strains from DFIs in Tunisia, showing a high genetic diversity, moderate antimicrobial resistance, but a high number of virulence-related traits, highlighting their pathological importance.

Association of HLA-G 3'UTR polymorphisms with hepatitis B virus infection in Tunisian population.

Hepatitis B virus (HBV) infection is a major public health burden. The mechanisms of immune evasion during chronic HBV (CHB) infection are poorly understood. Human leukocyte antigen (HLA)-G, an immune checkpoint molecule, plays a crucial role in the tolerance mechanisms of various infectious diseases. The 3' untranslated region (3'UTR), including the HLA-G + 3142 C > G polymorphism (rs1063320) and the 14-pb Ins/Del (rs66554220) has been strongly suggested to influence HLA-G expression. This study conducted a case-control analysis to evaluate the potential correlation between the HLA-G + 3142 C > G polymorphism and HBV infection outcome in a Tunisian cohort. The HLA-G + 3142 C > G polymorphism was analysed by PCR-RFLP in 242 patients with chronic HBV infection (116 males and 126 females), 241 healthy controls (116 males and 125 females), and 100 spontaneously resolved subjects (52 males and 48 females). Patients with chronic HBV infection showed a higher frequency of the + 3142G allele compared to healthy controls and spontaneously resolved subjects (p = 0.001 and p = 0.002, respectively). An association between the + 3142G allele and high HBV DNA levels was observed when HBV patients were stratified based on their HBV DNA levels (p = 0.016). Furthermore, the dominant model (GG + GC vs CC) was associated with liver function parameters, including AST, ALT, and high HBV DNA levels (p = 0.04, p < 0.001 and p = 0.002, respectively). However, there was no significant association found between this polymorphism and the fibrosis stage (p = 0.32). The haplotype analysis, using a subset of previously published data on the HLA-G 14-pb Ins/Del polymorphism, revealed an association between the Ins/G haplotype and chronic HBV infection (H1: InsG, p < 0.001). Our findings suggest that the + 3142G allele is a risk factor for the persistence and progression of HBV infection, while the + 3142C allele serves as a protective allele associated with the spontaneous resolution of the infection. Additionally, the HLA-G 3'UTR haplotype Ins/G is associated with chronic HBV infection in the Tunisian population.

Surveillance of Enterobacteriaceae from Diabetic Foot Infections in a Tunisian Hospital: Detection of E. coli-ST131-blaCTX-M-15 and K. pneumoniae-ST1-blaNDM-1 Strains.

The study determined the prevalence, antimicrobial resistant (AMR) determinants, and genetic characteristics of Escherichia coli and Klebsiella pneumoniae isolates from patients with diabetic foot infection (DFI) in a Tunisian hospital. A total of 26 Escherichia spp. and Klebsiella spp. isolates were recovered and identified by MALDI-TOF-MS. Antimicrobial susceptibility testing, the detection of AMR determinants and Shiga-like toxin genes, phylogenetic grouping, and molecular typing were performed. Twelve E. coli, 10 K. pneumoniae, 3 K. oxytoca, and 1 E. hermanii were isolated. A multidrug-resistant phenotype was detected in 65.4% of the isolates. About 30.8% of isolates were extended-spectrum ß-lactamase (ESBL) producers and mainly carried blaCTX-M-15 and blaCTX-M-14 genes. One blaNDM-1-producing K. pneumoniae-ST1 strain was identified. Class 1 integrons were detected in 11 isolates and 5 gene cassette arrangements were noted: dfrA1+aadA1 (n = 1), dfrA12+aadA2 (n = 3), and dfrA17+aadA5 (n = 1). Other non-ß-lactam resistance genes detected were as follows (number of isolates): aac(3')-II (3), aac(6')-Ib-cr(8), qnrB (2), qnrS (4), cmlA (2), floR (4), sul1 (11), sul2 (11), and sul3 (2). The phylogroup B1 was the most frequent (41.7%) among E. coli, and two ESBL-producing isolates corresponded to the ST131-B2 lineage. The ESBL- and carbapenemase-producing Enterobacteriaceae in DFIs are described for the first time in Tunisia.

Bioremediation of petroleum refinery wastewater by fungal stains isolated from the fishing harbour of Bizerte (Mediterranean Sea).

The study was conducted in order to explore the potential of fungi isolated from surface and bottom seawater collected from the fishing harbour of Bizerte on the bioremediation of industrial effluent (IE) contaminated by petroleum hydrocarbon. Among the 128 fungal isolates, 11 were isolated from surface seawater and 7 from bottom seawater, representing 18 taxa in total. The gas chromatography mass spectrometry (GC-MS) was used for the determination of hydrocarbon compounds in IE. An initial screening of fungal growth using six concentrations ranged between 20 and 70% (v/v) IE has allowed the identification of the optimal concentration for fungal growth as well as selection of species able to tolerate high amounts of hydrocarbon. Colorimetric test employing 2,6-dichlorophenol indophenol and gravimetric method was applied for the assessment of fungal growth using 20% EI. By checking the phylogenetic affiliation of the high-performing stains as identified using ITSr DNA sequence, a dominance of Ascomycetes was detected. Indeed, Aspergillus terreus and Penicillium expansum may degrade 82.07 and 81.76% of residual total petroleum hydrocarbon (TPH), respectively. Both species were collected from surface seawater. While, Aspergillus niger, Colletotrichum sp and Fusarium annulatum displayed comparable degradation rates 40.43%, 41.3%, and 42.03%, respectively. The lowest rate of degradation 33.62% was detected in Emericellopsis phycophila. All those species were isolated from bottom seawater, excepting A. niger isolated from surface water. This work highlighted the importance of exploring the potential of fungi isolated from the natural environment on the bioremediation of industrial effluent. Our results promoted the investigation of the potential of the high-performing isolates A. terreus and P. expansum on the bioremediation of IE at pilot-scale and then in situ.

Characterization of carbapenemase-producing Gram-negative bacilli: first report of blaNDM-1 in Enterobacter cloacae.

INTRODUCTION: The spread of multidrug-resistant bacteria, particularly carbapenem-resistant Gram-negative bacilli (CR-GNB), has become a serious challenge for clinicians due to limited therapeutic options. The aim of the study was to investigate the prevalence of carbapenemase production among clinical isolates recovered from 352 samples collected in Tebessa hospital, Algeria. METHODOLOGY: Bacterial isolates were identified by 16S RNA gene sequencing and susceptibility to antibiotics was determined by disk diffusion method. Carbapenem-resistant isolates were screened for carbapenemase production using modified carba Nordmann-Poirel test, modified Hodge test and imipenem-EDTA combined disc test. Extended-spectrum ß-lactamases (ESBL) were detected using double-disk synergy test. Molecular characterization of carbapenemases and ESBL genes was performed by polymerase chain reaction (PCR) and sequencing. RESULTS: A total of 85 Gram-negative bacilli isolates were recovered mainly from urine samples and were identified as: Klebsiella pneumoniae (17.65%), Serratia odorifera (15.29%), Escherichia coli (12.94%), Raoultella ornithinolytica, Enterobacter cloacae (11.76%), Serratia marcescens (10.59%), Morganella morganii (7.06%), Proteus mirabilis (5.88%), Acinetobacter baumannii (4.70%) and Pseudomonas aeruginosa (2.35%). All strains were resistant or intermediate to imipenem and/or ertapenem. ESBL, carbapenemase and metallo-beta-lactamases (MBL) phenotypes were detected in 19 (22.35%), 9 (10.59%) and 2 (2.35%) GNB isolates, respectively. PCR results in nine carbapenemase-producing GNB strains chosen showed the presence of one to four carbapenemase genes (blaGES, blaSME, blaNDM-1, blaVIM, blaGIM, blaSPM, blaOXA-48) in four strains; however, seven strains had at least one ESBL gene (blaTEM-1, blaCTXM-15, blaSHV). CONCLUSIONS: In this study, we report the first incidence of blaNDM-1 gene in Enterobacter cloacae isolated from urine sample in Algeria.

Isolation and characterization of a newly chrysene-degrading Achromobacter aegrifaciens.

Polycyclic aromatic hydrocarbons (PAHs) are considered substances of potential human health hazards because of their resistance to biodegradation and carcinogenic index. Chrysene is a PAH with a high molecular weight (HMW) that poses challenges for its elimination from the environment. However, bacterial degradation is an effective, environmentally friendly, and cost-effective solution. In our study, we isolated a potential chrysene-degrading bacteria from crude oil-contaminated seawater (Bizerte, Tunisia). Based on 16SrRNA analysis, the isolate S5 was identified as Achromobacter aegrifaciens. Furthermore, the results revealed that A. aegrifaciens S5 produced a biofilm on polystyrene at 20 °C and 30 °C, as well as at the air-liquid (A-L) interface. Moreover, this isolate was able to swim and produce biosurfactants with an emulsification activity (E24%) over 53%. Chrysene biodegradation by isolate S5 was clearly assessed by an increase in the total viable count. Confirmation was obtained via gas chromatography-mass spectrometry (GC-MS) analyses. A. aegrifaciens S5 could use chrysene as its sole carbon and energy source, exhibiting an 86% degradation of chrysene on day 7. In addition, the bacterial counts reached their highest level, over 25 × 1020 CFU/mL, under the conditions of pH 7.0, a temperature of 30 °C, and a rotary speed of 120 rpm. Based on our findings, A. aegrifaciens S5 can be a potential candidate for bioremediation in HMW-PAH-contaminated environments.

High Prevalence of GES-5 Variant and Co-Expression of VIM-2 and GES-45 among Clinical Pseudomonas aeruginosa Strains in Tunisia.

Carbapenem-resistant Pseudomonas aeruginosa (CRPA) are a global health concern. The antimicrobial resistance, virulence, and molecular typing of 57 CRPA isolated from 43 patients who attended a specific Tunisian hospital from September 2018 to July 2019 were analyzed. All but one were multidrug-resistant CRPA, and 77% were difficult-to-treat-resistant (DTR) isolates. The blaVIM-2 gene was detected in four strains (6.9%), and among the 36 blaGES-positive CRPA (62%), the blaGES-5 gene was the predominant variant (86%). Three strains co-harbored the blaVIM-2 and blaGES-45 genes, and seven CRPA carried the blaSHV-2a gene (14%). OprD alterations, including truncations by insertion sequences, were observed in 18 strains. Regarding the 46 class 1 integron-positive CRPA (81%), the blaGES-5 gene was located in integron In717, while the blaGES-29 and blaGES-45 genes were found in two new integrons (In2122 and In4879), and the blaVIM-2 gene was found in In1183 and the new integron In2142. Twenty-four PFGE patterns and thirteen sequence types (three new ones) were identified. The predominant serotype O:11 and exoU (81%) were mostly associated with ST235 and the new ST3385 clones. The seven blaSHV-2a-CRPA from different patients belonged to ST3385 and the same PFGE pattern. The blaGES-5- and blaVIM-2 + blaGES-45-positive CRPA recovered mostly from ICU patients belonged to the high-risk clone ST235. Our results highlight the alarming prevalence of blaGES-5- and ST235-CRPA, the co-existence of blaGES-45 and blaVIM-2, and their location within integrons favoring their dissemination.

Zinc Tetrafluoroborate-Mediated Ring Expansion of trans-Aziridine-2-carboxylates to cis-2-Iminothiazolidines and cis-Thiazolidine-2-iminium Tetrafluoroborates and Evaluation of Antimicrobial Activity.

Cis-2-iminothiazolidines and cis-thiazolidine-2-iminium tetrafluoroborates were successfully produced from trans-N-alkyl aziridine-2-carboxylates and phenyl/alkyl isothiocyanates mediated by zinc tetrafluoroborate in refluxing DCE. Reactions were performed via a complete regio- and stereoselective process to give the title iminothiazolidines and cis-thiazolidine-2-iminium salts in moderate to good yields (35 to 82%) with a wide substrate scope. In addition, the antibacterial activity evaluation of these compounds, as well as the minimum inhibitory concentration (MIC) determination, revealed that only four cis-thiazolidine-2-iminium salts showed growth inhibition against Bacillus cereus.

Physicochemical and Microbiological Water Quality Assessment of a Northwestern Algerian Dam: Detection of Ichtyopathogenic Bacteria.

Freshwater fish are often exposed to threats from anthropogenic or natural origins, such as pathogenic or opportunistic microorganisms responsible for a broad range of severe infections. In this study, we aimed to assess this microbiological threat to fish in an Algerian northwestern dam Sekkak (Tlemcen) by evaluating the diversity of ichtyopathogenic bacteria. In order to determine the water quality, physicochemical analyses of the dam water were carried out in situ. Ichtyopathogenic bacteria were isolated on selective media and identified by API galleries and molecular techniques (PCR and sequencing of the 16S rRNA gene). Besides, the antibiograms were constructed for all the isolates. The physicochemical and bacteriological analyses allowed us to classify the dam water as moderately polluted to polluted. Furthermore, an important diversity of ichtyopathogenic bacterial species was observed as Aeromonas hydrophila, Providencia rettgeri, and Pseudomonas aeruginosa were retrieved. The antibiogram test revealed notable resistance. The antibiotic family for which most resistances were found was the ß-lactam family, followed by aminoglycosides and macrolides. These results indicate that aquatic environments can shelter multidrug-resistant pathogenic bacteria representing a threat to the endemic fauna. Therefore, it is important to closely monitor these waters in order to improve the fish's living environment and ensure healthier production.

Reproductive Performance of Tunisian Arabian Stallions: A Study on the Variance and Estimation of Heritability.

A consistently high level of stallion fertility plays an economically important role in modern horse breeding. To better understand the factors affecting variation in stallion fertility, we have performed a statistical analysis study on some breeding factors: year of breeding, stud farm, age of the stallion, number of covered mares per stallion, reproduction methods, and age of the mare. This work was conducted on 94 purebred Arabian stallions in four different regions of Tunisia. The results showed an increase in the number of stallions during the study period, ranging from 11.33% in 2011 to 13.29% in 2018. Sidi Thabet's stud farm contained the highest number of purebred Arabian stallions. The majority of stallions were between 15 and 21 years old and had covered 1 to 20 mares; 95.19% of stallions were used in natural mating (Nat); 50.36% had low fertility, 17.69% had medium fertility, and 32.3% had excellent fertility according to fertility standards. Depending on the year and stud, there was a variation in fertility per cycle (FERPCE) and end-of-season fertility (FERPSE) of the stallions. The highest average FERPCE and FERPSE values were obtained using artificial insemination with fresh semen (AIF). Analysis of FERPCE and FERPSE showed that the model used in our study explained 40.21% of total variability observations for FERPCE and 42.1% for FERPSE. The used statistical model showed that the breeding year, the stud, the age of the stallion, the number of covered mares by stallions and the method of reproduction significantly affected both FERPCE and FERPSE (with p = 0.001). Low to moderate heritability estimations for FERPCE (hs2 = 0.08) and FERPSE (hes2 = 0.36) were obtained.

Antimicrobial Activities and Mode of Flavonoid Actions.

The emergence of antibiotics-resistant bacteria has been a serious concern for medical professionals over the last decade. Therefore, developing new and effective antimicrobials with modified or different modes of action is a continuing imperative. In this context, our study focuses on evaluating the antimicrobial activity of different chemically synthesized flavonoids (FLAV) to guide the chemical synthesis of effective antimicrobial molecules. A set of 12 synthesized molecules (4 chalcones, 4 flavones and 4 flavanones), bearing substitutions with chlorine and bromine groups at the C6' position and methoxy group at the C4' position of the B-ring were evaluated for antimicrobial activity toward 9 strains of Gram-positive and Gram-negative bacteria and 3 fungal strains. Our findings showed that most tested FLAV exhibited moderate to high antibacterial activity, particularly against Staphylococcus aureus with minimum inhibitory concentrations (MIC) between the range of 31.25 and 125 µg/mL and that chalcones were more efficient than flavones and flavanones. The examined compounds were also active against the tested fungi with a strong structure-activity relationship (SAR). Interestingly, leakage measurements of the absorbent material at 260 nm and scanning electron microscopy (SEM) demonstrated that the brominated chalcone induced a significant membrane permeabilization of S. aureus.

Comparative genomics reveals the acquisition of mobile genetic elements by the plant growth-promoting Pantoea eucrina OB49 in polluted environments.

Heavy metal-tolerant plant growth-promoting bacteria (PGPB) have gained popularity in bioremediation in recent years. A genome-assisted study of a heavy metal-tolerant PGPB Pantoea eucrina OB49 isolated from the rhizosphere of wheat grown on a heavy metal-contaminated site is presented. Comparative pan-genome analysis indicated that OB49 acquired heavy metal resistance genes through horizontal gene transfer. On contigs S10 and S12, OB49 has two arsRBCH operons that give arsenic resistance. On the S12 contig, an arsRBCH operon was discovered in conjunction with the merRTPCADE operon, which provides mercury resistance. P. eucrina OB49 may be involved in an ecological alternative for heavy metal remediation and growth promotion of wheat grown in metal-polluted soils. Our results suggested the detection of mobile genetic elements that harbour the ars operon and the fluoride resistance genes adjacent to the mer operon.

Physiological and genomic insights into abiotic stress of halophilic archaeon Natrinema altunense 4.1R isolated from a saline ecosystem of Tunisian desert.

Halophilic archaea are polyextremophiles with the ability to withstand fluctuations in salinity, high levels of ultraviolet radiation, and oxidative stress, allowing them to survive in a wide range of environments and making them an excellent model for astrobiological research. Natrinema altunense 4.1R is a halophilic archaeon isolated from the endorheic saline lake systems, Sebkhas, located in arid and semi-arid regions of Tunisia. It is an ecosystem characterized by periodic flooding from subsurface groundwater and fluctuating salinities. Here, we assess the physiological responses and genomic characterization of N. altunense 4.1R to UV-C radiation, as well as osmotic and oxidative stresses. Results showed that the 4.1R strain is able to survive up to 36% of salinity, up to 180 J/m2 to UV-C radiation, and at 50 mM of H2O2, a resistance profile similar to Halobacterium salinarum, a strain often used as UV-C resistant model. In order to understand the genetic determinants of N. altunense 4.1R survival strategy, we sequenced and analyzed its genome. Results showed multiple gene copies of osmotic stress, oxidative stress, and DNA repair response mechanisms supporting its survivability at extreme salinities and radiations. Indeed, the 3D molecular structures of seven proteins related to responses to UV-C radiation (excinucleases UvrA, UvrB, and UvrC, and photolyase), saline stress (trehalose-6-phosphate synthase OtsA and trehalose-phosphatase OtsB), and oxidative stress (superoxide dismutase SOD) were constructed by homology modeling. This study extends the abiotic stress range for the species N. altunense and adds to the repertoire of UV and oxidative stress resistance genes generally known from haloarchaeon.

First Report of IMI-2-Producing Enterobacter bugandensis and CTX-M-55-Producing Escherichia coli isolated from Healthy Volunteers in Tunisia.

The aim of this study was to characterize the prevalence of fecal carriage of extended-spectrum beta-lactamases and carbapenemase-producing Gram-negative bacteria among healthy humans in Tunisia. Fifty-one rectal swabs of healthy volunteers were plated on MacConkey agar plates supplemented with cefotaxime or imipenem. The occurrences of resistance genes, integrons, and phylogroup typing were investigated using PCR and sequencing. The genetic relatedness of isolates was determined by pulsed-field-gel-electrophoresis (PFGE) and multilocus-sequence-typing (MLST). Whole-genome-sequencing (WGS) was performed for the carbapenem-resistant isolate. Sixteen ESBL-producing Escherichia coli isolates and one carbapenem-resistant Enterobacter bugandensis were detected out of the fifty-one fecal samples. The ESBL-producing E. coli strains contained genes encoding CTX-M-15 (n = 9), CTX-M-1 (n = 3), CTX-M-27 (n = 3), and CTX-M-55 (n = 1). Three CTX-M-1-producers were of lineages ST131, ST7366, and ST1158; two CTX-M-15-producers belonged to lineage ST925 and ST5100; one CTX-M-27-producer belonged to ST2887, and one CTX-M-15-producer belonged to ST744. Six isolates contained class 1 integrons with the following four gene cassette arrangements: dfrA5 (two isolates), dfrA12-orf-aadA2 (two isolates), dfrA17-aadA5 (one isolate), and aadA1 (one isolate). E. bugandensis belonged to ST1095, produced IMI-2 carbapenemase, and contained qnrE1 and fosA genes. A genome-sequence analysis of the E. bugandensis strain revealed new mutations in the blaACT and qnr genes. Our results reveal an alarming rate of ESBL-E. coli in healthy humans in Tunisia and the first description of IMI-2 in E. bugandensis.

Co-occurrence of antibiotic and metal resistance in long-term sewage sludge-amended soils: influence of application rates and pedo-climatic conditions.

Urban sewage sludge (USS) is increasingly being used as an alternative organic amendment in agriculture. Because USS originates mostly from human excreta, partially metabolized pharmaceuticals have also been considered in risk assessment studies after reuse. In this regard, we investigated the cumulative effect of five annual USS applications on the spread of antibiotic-resistant bacteria (ARB) and their subsequent resistance to toxic metals in two unvegetated soils. Eventually, USS contained bacterial strains resistant to all addressed antibiotics with indices of resistance varying between 0.25 for gentamicin to 38% for ampicillin and azithromycin. Sludge-amended soils showed also the emergence of resistome for all tested antibiotics compared to non-treated controls. In this regard, the increase of sludge dose generally correlated with ARB counts, while soil texture had no influence. On the other hand, the multi-antibiotic resistance (MAR) of 52 isolates selected from USS and different soil treatments was investigated for 10 most prescribed antibiotics. Nine isolates showed significant MAR index (≥ 0.3) and co-resistance to Cd, As and Be as well. However, events including an extreme flash flood and the termination of USS applications significantly disrupted ARB communities in all soil treatments. In any case, this study highlighted the risks of ARB spread in sludge-amended soils and a greater concern with the recent exacerbation of antibiotic overuse following COVID-19 outbreak.

Methicillin-Resistant Staphylococcus aureus from Diabetic Foot Infections in a Tunisian Hospital with the First Detection of MSSA CC398-t571.

This study sought to analyze the antimicrobial resistant phenotypes and genotypes as well as the virulence content of S. aureus isolates recovered from patients with diabetic foot infections (DFIs) in a Tunisian hospital. Eighty-three clinical samples of 64 patients were analyzed, and bacterial isolates were identified by MALDI-TOF. The antimicrobial resistance phenotypes were determined by the Kirby-Bauer disk diffusion susceptibility test. Resistance and virulence genes, agr profile, spa and SCCmec types were determined by PCR and sequencing. S. aureus was detected in 14 of the 64 patients (21.9%), and 15 S. aureus isolates were recovered. Six out of the fifteen S. aureus isolates were methicillin-resistant (MRSA, mecA-positive) (40%). The isolates harbored the following resistance genes (number of isolates): blaZ (12), erm(B) (2), erm(A) (1), msrA (2), tet(M) (2), tet(K) (3), tet(L) (1), aac(6')-aph(2″) (2), ant(4″) (1) and fexA (1). The lukS/F-PV and tst genes were detected in three isolates. Twelve different spa-types were identified and assigned to seven clonal complexes with the predominance of agr-type III. Furthermore, the SCCmec types III, IV and V were found among the MRSA isolates. Moreover, one MSSA CC398-t571-agr-III isolate was found; it was susceptible to all antimicrobial agents and lacked luk-S/F-PV, tst, eta and etb genes. This is the first report on the prevalence and molecular characterization of S. aureus from DFIs and also the first detection of the MSSA-CC398-t571 clone in human infections in Tunisia. Our findings indicated a high prevalence S. aureus in DFIs with genetic diversity among the MSSA and MRSA isolates.

Diversity and adaptation properties of actinobacteria associated with Tunisian stone ruins.

Stone surface is a unique biological niche that may host a rich microbial diversity. The exploration of the biodiversity of the stone microbiome represents a major challenge and an opportunity to characterize new strains equipped with valuable biological activity. Here, we explored the diversity and adaptation strategies of total bacterial communities associated with Roman stone ruins in Tunisia by considering the effects of geo-climatic regions and stone geochemistry. Environmental 16S rRNA gene amplicon was performed on DNA extracted from stones samples collected in three different sampling sites in Tunisia, along an almost 400km aridity transect, encompassing Mediterranean, semiarid and arid climates. The library was sequenced on an Illumina MiSeq sequencing platform. The cultivable Actinobacteria were isolated from stones samples using the dilution plate technique. A total of 71 strains were isolated and identified based on 16S rRNA gene sequences. Cultivable actinobacteria were further investigated to evaluate the adaptative strategies adopted to survive in/on stones. Amplicon sequencing showed that stone ruins bacterial communities were consistently dominated by Cyanobacteria, followed by Proteobacteria and Actinobacteria along the aridity gradient. However, the relative abundance of the bacterial community components changed according to the geo-climatic origin. Stone geochemistry, particularly the availability of magnesium, chromium, and copper, also influenced the bacterial communities' diversity. Cultivable actinobacteria were further investigated to evaluate the adaptative strategies adopted to survive in/on stones. All the cultivated bacteria belonged to the Actinobacteria class, and the most abundant genera were Streptomyces, Kocuria and Arthrobacter. They were able to tolerate high temperatures (up to 45°C) and salt accumulation, and they produced enzymes involved in nutrients' solubilization, such as phosphatase, amylase, protease, chitinase, and cellulase. Actinobacteria members also had an important role in the co-occurrence interactions among bacteria, favoring the community interactome and stabilization. Our findings provide new insights into actinobacteria's diversity, adaptation, and role within the microbiome associated with stone ruins.

Meta-analysis of HLA-G 14bp insertion/deletion polymorphism and soluble HLA-G revealed an association with digestive cancers initiation and prognosis.

Background/Objective: Conflicting results on the association between HLA-G and digestive cancers were reported. We conducted a meta-analysis to further investigate the true relationship between HLA-G and digestive cancers (DC). Methods: Following PRISMA guidelines, we performed a meta-analysis including 7 case-control studies on HLA-G 14-bp Insertion/deletion (I/D) polymorphism, and 15 studies on soluble HLA-G (sHLA-G). Odds ratios (OR) and their corresponding 95% confidence intervals (CI) for genetic polymorphisms were calculated. The pooled OR was calculated under three genetic models: allelic, recessive, and dominant models. Concerning sHLA-G meta-analysis, standardized mean differences (SMDs) were calculated. Results: The HLA-G 14-bp I/D was not associated with the risk of DC. However, in the subset of HBV/HCV positive hepato-cellular cancer (HCC) patients, we reported a significant association of HLA-G 14-bp I/D with the disease initiation under allelic (D vs. I; OR = 1.698, 95% CI = 1.263-2.282, p = 0.000), dominant (DD + ID vs. II; OR = 2.321, 95% CI = 1.277-4.218, p = 0.006)and recessive (DD vs. DI + II; OR = 1.739, 95% CI = 1.173-2.577, p = 0.006) genetic models. Interestingly, HLA-G 14-bp I/D was not associated with the disease initiation in HBV/HCV negative HCC patients. However, the infection by HBV/HCV seems to be implicated in the HCC development when we compared HBV/HCV positive patients to HBV/HCV negative patients under allelic (D vs. I; OR = 1.429, 95% CI = 1.029-1.983, p = 0.033, and dominant (DD + ID vs.II; OR = 1.981, 95% CI = 1.002-3.916, p = 0.049) genetic models.Overall analysis of DC showed significant increased sHLA-G in patients compared to healthy controls (SMD = 3.341, 95% CI = 2.415-4.267, p = 0.000). In Asian patients with gastric cancer, sHLA-G was significantly increased in grade 3 compared to low grades (SMD = 0.448, 95% CI = 0.109-0.787, p = 0.000). Further analysis showed that sHLA-G was significantly increased in positive DC vascular invasion (SMD = 0.743, 95% CI = 0.385-1.100, p = 0.000). Accordingly, sHLA-G was associated with a poor prognosis for DC. Conclusion: The current meta-analysis supports the significant role of HLA-G in DC. The HLA-G 14-bp I/D polymorphism was associated with HCC patients with concomitant HBV/HCV viral infections. Increased sHLA-G indicated a poor prognosis for DC cancer patients.

The relationship of 3'UTR HLA-G14-bp insertion/deletion and +3142 C/G polymorphisms and soluble HLA-G expression with gynecological cancers: An updated meta-analysis.

OBJECTIVES: Human leukocyte antigen-G (HLA-G) is implicated in several cancers and is considered to be an immune checkpoint regulator. We determined the association between polymorphisms in the 3' untranslated region of HLA-G and soluble HLA-G (sHLA-G) expression with gynecological cancers (GCs). METHODS: A meta-analysis was conducted to examine the association between HLA-G14-bp insertion (I)/deletion (D) and +3142C/G polymorphism in GC and to evaluate sHLA-G expression RESULTS: We revealed a significant association between the +3142C/G polymorphism and invasive cervical cancer (ICC) based on the allelic model G versus C (odds ratio [OR] = 0.738, 95% confidence interval [CI] = 0.563-0.966, p = 0.027), dominant GG+GC versus CC (OR = 0.584, 95% CI = 0.395-0.862, p = 0.007), and codominant GG versus CC (OR = 0.527, 95% CI = 0.312-0.891, p = 0.017) models, suggesting that the G allele and GG genotype are protective against ICC. In gynecological precancerous patients with human papillomavirus (HPV) infection, we found that the 14-bp I/D under the codominant DD versus DI model (OR = 0.492, 95% CI = 0.241-1.004, p = 0.051) was of borderline significance. Soluble HLA-G levels were significantly higher in patients compared with healthy controls (standardized mean differences [SMD] = 1.434, 95% CI = 0.442-2.526, p = 0.005). Stratification by cancer type revealed that the sHLA-G levels were significantly increased in cervical cancer (SMD = 4.889, 95% CI = 0.468-9.310, p = 0.030) and in subjects of Asian ethnicity (SMD = 4.889, 95% CI = 0.467-9.309, p = 0.030). CONCLUSIONS: HLA-G14-bp I/D and +3142 C/G polymorphisms are associated with GC and HPV-associated cervical cancer. In addition, we found significantly increased sHLA-G levels in cancer patients. These results provide a basis for further studies in diagnostics and immunotherapy of GC.

Evaluation of the Potency of the Pertussis Vaccine in Experimental Infection Model with Bordetella pertussis: Study of the Case of the Pertussis Vaccine Used in the Expanded Vaccination Program in Algeria.

In Algeria, vaccination against pertussis is carried out using the whole-cell pertussis vaccine combined with the diphtheria and tetanus toxoids (DTwp). The quality control of vaccines locally produced or imported is carried out before the batch release. The aim of our work was to evaluate the potency of pertussis vaccines. In the present study, five consecutive trials of potency were conducted on samples of the same batch of (DTwp) using the mouse protection test (MPT) against experimental infection of Bordetella pertussis strain 18323, based on the Kendrick test. The virulence of B. pertussis strain 18-323 was verified by the mortality of mice, with an average LD50 of 338.92, as well as the dose of the lethal test containing a mean number of LD50 of 324.43. The (MPT) test recorded a relative potency of 8.02 IU/human dose, with 95% CL of (3.56-18.05) IU/human dose. The development of the (MPT) at the laboratory of quality control of vaccines and sera at the Pasteur Institute of Algeria was effective in evaluating the potency of whole-cell pertussis vaccines. Interestingly, our study indicates that this potency is necessary for the vaccine quality assurance. Further validation is needed to strengthen the application and routine use of the test.