Cross-sectional Molecular Detection of Francisella tularensis in Domestic Rabbits in Sulaimani Province Kurdistan Region, Iraq

Tularemia is one of the diseases transmitted between humans and animals. It is caused by a Gram-negative bacterium Francisella tularensis. Recent serological studies suggested that tularemia can be an endemic bacterial zoonotic disease in some countries surrounding Iraq such as Iran and Turkey. The main objective of this study is to detect tularemia in Sulaimani province northeast Iraq near to Iran border. Sulaimani city also has contact with many Turkish cities. This study was conducted between Jun and October 2020. Blood samples were taken from one hundred local breed rabbits of different ages and sexes. A highly sensitive real-time PCR technique was used. Sixteen out of one hundred blood samples (16%) were positively taken from different local breed rabbits from four different places in Sulaimani province. All positive samples were detected in the center of Sulaimani city. No published documents have been reported yet about tularemia in Kurdistan Region. This paper documented molecular detection of F. tularensis in local breed rabbits in Sulaimani province Kurdistan Region-Iraq

An investigation of the effect of the extracts from the seeds of Bidens pilosa on the growth of Escherichia coli and Bacillus subtilis

Escherichia coli is a Gram-negative bacterium also found in the human. Bacillus subtilis is a Gram-positive, non-pathogenic, endospore-forming bacterium. The spores can survive the heat and B. subtilis to cause food poisoning. The study hypothesized that the extracts from B. pilosa would inhibit the growth of E. coli and B. subtilis. The seeds of B. pilosa were purchased from the seed company. The seeds were dried, ground, and shaken in acetone, ethanol, methanol, and water for 72 hours. Solvents were evaporated and the crude extracts were used for antibacterial activity using a modified Kirby-Bauer disc method. The results revealed that the growth of E. coli was inhibited by the extracts using ethanol. The zones of inhibition were 13 mm. The extracts that were extracted using acetone and water were not effective in inhibiting the growth of E. coli. The growth of B. subtilis was inhibited by the extracts from acetone and methanol. The zones of inhibition were 13 mm. The results revealed that the extracts from ethanol and water were not effective in inhibiting the growth of B. subtilis. Seeds of B. pilosa have the potential to be used as antimicrobials.

Multilocus Sequence Typing of Non-JP2 Serotype b Aggregatibacter actinomycetemcomitans Strains of Ghanaian and Swedish Origin

Objective and MethodsThe Gram-negative bacterium, Aggregatibacter actinomycetemcomitans is associated with periodontitis affecting young individuals. The geographic dissemination of the highly leukotoxic JP2 genotype of serotype b of this species was previously studied by multilocus sequence typing (MLST). Here, we have used MLST to genetically characterize non-JP2 genotype strains of serotype b, isolated from individuals living in Ghana (n=41), and in Sweden (n=13), respectively.ResultsThe MLST analysis revealed a total of nine sequence types (ST). Both Ghanaian and Swedish isolates were distributed in ST 1-3. ST 5 and 6 were only identified among the Ghanaian strains, whereas ST 4, 7, 8 and 9 were uniquely represented among the Swedish strains. Previously, we characterized these non-JP2 genotype strains of A. actinomycetemcomitans serotype b by arbitrarily-primed (AP)-PCR, which distributed them into three groups, AP-PCR type 1, 2, and 3, respectively. AP-PCR type 1 strains are generally highly leukotoxic, and are associated with progression of periodontal attachment loss. As AP-PCR type 1 includes both JP2 genotype strains and a proportion of non-JP2 genotype strains of serotype b, a straightforward diagnostic procedure has been sought. This has revealed a gene, cagE, which appears to be conserved only in this AP-PCR type. According to our results, MLST was not a highly discriminatory method to identify AP-PCR type 1, as strains of this AP-PCR type could be found within three different ST: ST 2, ST 3 and ST 8.ConclusionAccording to MLST, a geographic dissemination of non-JP2 genotype A. actinomycetemcomitans serotype b appears to exist. However, aiming to identify carriers of AP-PCR type 1, non-JP2 genotype serotype b, PCR with cagE-specific primers is likely the most efficient diagnostic procedure known today.

The Effect of Raw Materials on the Properties of Oxidized Cellulose Nanofibers from Bacterial Cellulose

In the present study, nanofibers of oxidized cellulose (OC) were prepared from dried bacterial cellulose using a mixture of nitric acid/phosphoric acid and sodium nitrite. Three types of dried bacterial cellulose were used as raw materials. The results revealed that dried sheet bacterial cellulose (DSBC) yielded 86.8% oxidized cellulose with 19.4% carboxyl content, whereas squeeze-dried bacterial cellulose (SDBC) yielded 53.3% OC with 28.6% carboxyl content, and freeze-dried bacterial cellulose (FDBC) yielded 55.6% of OC with 27.6% carboxyl content. The results revealed that OC neutralized with sodium hydroxide from SDBC showed the best swelling property among all types of OC. SDBC indicated the reduction of CFU exceeds 99.99% for gram-negative bacterium Escherichia coli ATCC 25922 and gram-positive bacterium Staphylococcus aureus ATCC 6538.

CircNFIC Balances Inflammation and Apoptosis by Sponging miR-30e-3p and Regulating DENND1B Expression

Disordered inflammation and apoptosis are closely related to diseases, and inflammation can also promote cell apoptosis, where growing evidence has shown that circular RNAs (circRNAs) play important roles. Lipopolysaccharide (LPS) is the main component of the cytoderm of gram-negative bacterium, which can cause inflammatory responses in macrophages. We constructed an inflammatory model by exposing chicken macrophage cell lines (also known as HD11) to LPS for in vitro experiments. In this study, we validated a novel circRNA—circNFIC—which was dramatically up-regulated in tissues infected by coccidia and cells exposed to LPS. Besides, circNFIC could significantly promote the expression levels of pro-inflammation factors, including (IL-1β, TNFα, and IFNγ) and pro-apoptosis maker genes (caspase 3 and caspase 8) in HD11 exposed to LPS or not. In terms of mechanism, circNFIC exerted notable effects on DENND1B to regulate cell inflammation and apoptosis by sponging miR-30e-3p. The molecular functions played by miR-30e-3p and DENND1B have been explored, respectively. In addition, the effects of circNFIC knockdown suppressing the expression of pro-inflammatory and pro-apoptosis functions could be reversed by a miR-30e-3p inhibitor. On the whole, circNFIC promoted cell inflammation and apoptosis via the miR-30e-3p/DENND1B axis.

Hafnia alvei Pneumonia: A Rare Cause of Infection in a Patient with COVID-19

Herein, we describe a case report of a critically ill patient, a 48-year-old man without comorbidities admitted to the hospital with a serious type 1 (hypoxemic) respiratory insufficiency and confirmed diagnosis of COVID-19. After 5 days with invasive mechanical ventilation, the patient developed a bacterial co-infection, namely a pneumonia by Hafnia alvei, requiring the last line of respiratory support: extracorporeal membrane oxygenation (ECMO). Subsequently, his clinical situation gradually stabilized, until he was discharged from the hospital on day 61, being accompanied in ambulatory consultation by the physical medicine and pulmonology department during the post-COVID-19 recovery. H. alvei is a Gram-negative bacterium that is rarely isolated from human specimens and is rarely considered to be pathogenic. However, COVID-19 disease can cause substantial organ dysfunction and can be associated with bacterial secondary infections which can favor the emergence of rare infectious diseases by uncommon microorganisms.

Antimicrobial Peptides With Antibiofilm Activity Against Xylella fastidiosa

Xylella fastidiosa is a plant pathogen that was recently introduced in Europe and is causing havoc to its agriculture. This Gram-negative bacterium invades the host xylem, multiplies, and forms biofilm occluding the vessels and killing its host. In spite of the great research effort, there is no method that effectively prevents or cures hosts from infections. The main control strategies up to now are eradication, vector control, and pathogen-free plant material. Antimicrobial peptides have arisen as promising candidates to combat this bacterium due to their broad spectrum of activity and low environmental impact. In this work, peptides previously reported in the literature and newly designed analogs were studied for its bactericidal and antibiofilm activity against X. fastidiosa. Also, their hemolytic activity and effect on tobacco leaves when infiltrated were determined. To assess the activity of peptides, the strain IVIA 5387.2 with moderate growth, able to produce biofilm and susceptible to antimicrobial peptides, was selected among six representative strains found in the Mediterranean area (DD1, CFBP 8173, Temecula, IVIA 5387.2, IVIA 5770, and IVIA 5901.2). Two interesting groups of peptides were identified with bactericidal and/or antibiofilm activity and low-moderate toxicity. The peptides 1036 and RIJK2 with dual (bactericidal–antibiofilm) activity against the pathogen and moderate toxicity stand out as the best candidates to control X. fastidiosa diseases. Nevertheless, peptides with only antibiofilm activity and low toxicity are also promising agents as they could prevent the occlusion of xylem vessels caused by the pathogen. The present work contributes to provide novel compounds with antimicrobial and antibiofilm activity that could lead to the development of new treatments against diseases caused by X. fastidiosa.

Synthesis of Aminooxy Glycoside Derivatives of the Outer Core Domain of Pseudomonas aeruginosa Lipopolysaccharide

Pseudomonas aeruginosa is a highly prevalent gram-negative bacterium that is becoming more difficult to treat because of increasing antibiotic resistance. As chemotherapeutic treatment options diminish, there is an increased need for vaccines. However, the creation of an effective P. aeruginosa vaccine has been elusive despite intensive efforts. Thus, new paradigms for vaccine antigens should be explored to develop effective vaccines. In these studies, we have focused on the synthesis of two L-rhamnose–bearing epitopes common to glycoforms I and II of the outer core domain of Pseudomonas aeruginosa lipopolysaccharide, α-L-Rha-(1→6)-α-D-Glc-(1→4)-α-D-GalN-(Ala)-α-aminooxy (3) and α-L-Rha-(1→3)-β-D-Glc-(1→3)-α-D-GalN-(Ala)-α-aminooxy (4), respectively. The target trisaccharides were both prepared starting from a suitably protected galactosamine glycoside, followed by successive deprotection and glycosylation with suitably protected D-glucose and L-rhamnose thioglycosides. Global deprotection resulted in the formation of targets 3 and 4 in 22 and 35% yield each. Care was required to modify basic reaction conditions to avoid early deprotection of the N-oxysuccinamido group. In summary, trisaccharides related to the L-rhamnose–bearing epitopes common to glycoforms I and II of the outer core domain of Pseudomonas aeruginosa lipopolysaccharide have been prepared as their aminooxy glycosides. The latter are expected to be useful in chemoselective oxime-based bioconjugation reactions to form Pseudomonas aeruginosa vaccines.

UHPLC-ESI-Orbitrap-HR-MS Analysis of Cyclopeptide Alkaloids From Ziziphus joazeiro

Ziziphus joazeiro Mart., popularly known as “juazeiro”, is a tree widely found in the northeast of Brazil. It is commonly used as an anti-inflammatory, antibacterial, antifungal, and analgesic agent. The stem extract exhibited, beside cytotoxic properties, substantial activity against the Gram-negative bacterium Allivibrio fischeri. UHPLC-ESI-Orbitrap-HR-MS analysis of the alkaloidal fraction of the crude methanolic stem extract of this species enabled the detection and putative identification of sixteen cyclopeptide alkaloids (CPAs), including four possibly new structures. According to the MS2 fragmentation analysis, from the sixteen identified CPAs, three possess a type-Ia1, one a type-Ia2, and twelve a type-Ib cyclopeptide alkaloid core. The structures of paliurine-C and -D were supported by NMR data.

1009. Biofilm Formation in Acinetobacter Baumannii Clinical Isolates

Background Multidrug resistant Acinetobacter baumannii (MDR-Ab) is a Gram-negative bacterium known for causing severe nosocomial infections, attributed in part to its formation of biofilm. Siderophore is a virulence factor known to support biofilm formation by regulating iron availability. In this study, we screened 44 isolates of MDR-Ab from our Gram-negative repository to determine the strains that phenotypically form biofilm and produce siderophore. The results were compared to Pseudomonas aeruginosa PAO1, which produces both biofilm and siderophore. Methods Isolates were grown overnight in minimal M9 medium supplemented with casamino acids and hydroxyquinones at 37°C. Bacterial cells were normalized (to OD 600=0.01) and a standard diluted 10-3 tube was used in the study. A 96-well plate was inoculated with 100 microliters of each isolate in quadruplicates. This process was repeated in Tygon tubes with 50 microliters of each isolate in triplicates. The plate and Tygon tubes were incubated statically for 48 hours at 30°C and then stained with crystal violet. The contents were dissolved in 33% glacial acetic acid and analyzed by spectrophotometry to measure biofilm formation. Siderophore secretion was measured in supernatants with Chrome Azurol S (CAS) reagent and production was observed on CAS agar plates. Results High levels of biofilm formation were observed in 8 strains of MDR-Ab in the 96-well plate (3, 4, 9, 22, 61, 1010, 1012, 1022) and 6 strains in Tygon tubes (3, 4, 16, 66, 1002, 1010) (Fig. 1). There was minimal siderophore production in MDR-Ab isolates compared to PAO1 in both the 96-well plate and Tygon tubes (Fig. 2). Only 4 strains lacked siderophore production on CAS agar and were inversely negative for the secretion in medium. Figure 1 Biofilm formation in a 96-well plate and Tygon tubes (A) High levels of biofilm formation were observed in MDR-Ab strain numbers 3, 4, 9, 22, 61, 1010, 1012, 1022 in the 96-well plate. (B) High levels of biofilm formation were observed in MDR-Ab strain numbers 3, 4, 16, 66, 1002, 1010 in Tygon tubes. Figure 2 Degree of siderophore production in a 96-well plate and Tygon tubes Siderophore production of MDR-Ab was limited compared to PAO1 after inoculation in a 96-well plate (A) and in Tygon tubes (B). Conclusion Many strains of MDR-Ab readily form biofilm. Overall siderophore production is lower in MDR-Ab compared to consistent production by PAO1, but this does not appear to affect MDR-Ab’s ability to form biofilm. Unlike in PAO1, biofilm formation in MDR-Ab may occur independently of siderophore production. This research serves as a basis for understanding future MDR-Ab biofilm elimination in patient catheters and indwelling devices. Disclosures All Authors: No reported disclosures