Molecular Identification Methods Research Articles

A LAMP Assay for the Detection of Thecodiplosis japonensis, an Alien Gall Midge Species Pest of Pine Trees.

Simple SummaryThecodiplosis japonensis is considered the most harmful pest to pines in South Korea. T. japonensis is a native species of Japan. Recently, T. japonensis was discovered in China and has caused serious damage to local pine trees. Due to the small size and little morphological difference with its related species, it is difficult to accurately identify T. japonensis by morphological methods. Accurate and efficient molecular identification methods are urgently needed to detect this invasive gall midge pest, yet there was no molecular identification method for T. japonensis. In this study, we developed a LAMP assay to detect T. japonensis based on the COI gene sequence. The LAMP assay could detect as little as 300 fg of gDNA. Using colorimetric amplification and a crude gDNA extraction method, the total procedure could be processed in 75 min. The method established in the study can be easily used in both laboratory and field conditions, enabling rapid molecular identification of T. japonensis.Pine needle gall midge (T. japonensis), native to Japan, has become a serious invasive pest in South Korea and, more recently in 2006, in China. It was first discovered in Qingdao, Shandong Province, and has caused serious damage to local Pinus thunbergii. The insect’s small size makes morphological-based identification difficult; therefore, molecular detection techniques are urgently needed for monitoring and preventing its further spread. At present, there is no simple and accurate field molecular identification tool. To solve this problem, a LAMP-based molecular diagnosis technology of T. japonensis was developed. Four LAMP primers were designed to specifically amplify T. japonensis DNA. Positive LAMP reactions usually produce amplification in one hour. The optimal incubation conditions for LAMP detection were determined with 4 LAMP primers for 60 min at 61 °C. The LAMP detection range of gDNA concentrations is wide, with a minimum detectable gDNA concentration of 300 fg. A non-destructive DNA-releasing procedure, HotSHOT “HS6”, which could extract “crude DNA” for LAMP assay in 10 min, was used for larval and adult samples. Therefore, we established a LAMP-based rapid molecular identification method that can be applied in the monitoring and management of T. japonensis.

Molecular identification of the strains of the domestic silkworm, Bombyx mori (Lepidoptera: Bombycidae), which are endemic to Korea, based on single nucleotide polymorphisms in mitochondrial genome sequences

• We developed molecular identification methods for three endemic silkworm strains. • We found 15 SNP in mitogenomes of 5 endemic and 34 stock strains. • We applied the RFLP-PCR and the tetra-primer ARMS-PCR methods. • We distinguished three endemic strains using both methods in agarose gels. • These methods could be useful for distinguishing endemic strains preserved in Korea. The domestic silkworm, Bombyx mori (Lepidoptera: Bombycidae), has been diversified into various strains over a long period. However, methods to distinguish silkworm strains remain limited partially owing to the genetic similarity caused by the long history of domestication. In this study, we developed molecular identification methods to distinguish three domestic silkworm strains, which are endemic to Korea. By comparing publicly available complete mitochondrial genome (mitogenome) sequences of five endemic strains and 34 stock silkworm strains analyzed in a previous study, we detected 15 single nucleotide polymorphisms (SNPs; SNP1–SNP15), which distinguished the following three endemic strains: Sun7ho (SN7), Sandongsammyeon (SDS), and Sammyeonhonghoeback (SMH). We used two SNPs for each strain to identify the three endemic strains. To distinguish each SN7 and SDS from the remaining four endemic and 34 stock strains, the PCR-restriction fragment length polymorphism method was employed using Acu I and Hpa I restriction enzymes, which recognize SNP1 and SNP8, respectively. Additionally, the tetra-primer amplification refractory mutation system PCR method was used to determine the regions containing SNP3, SNP11, and both SNP14 and SNP15 to distinguish SN7, SDS, and SMH, respectively, from the remaining strains. A validation test with additional individuals showed that each target strain was clearly recognized, suggesting that mitogenome SNP-based methods can be used to identify three endemic silkworm strains during culture and breeding.

The Threat Called Candida haemulonii Species Complex in Rio de Janeiro State, Brazil: Focus on Antifungal Resistance and Virulence Attributes.

Although considered rare, the emergent Candida haemulonii species complex, formed by C. haemulonii sensu stricto (Ch), C. duobushaemulonii (Cd) and C. haemulonii var. vulnera (Chv), is highlighted due to its profile of increased resistance to the available antifungal drugs. In the present work, 25 clinical isolates, recovered from human infections during 2011–2020 and biochemically identified by automated system as C. haemulonii, were initially assessed by molecular methods (amplification and sequencing of ITS1-5.8S-ITS2 gene) for precise species identification. Subsequently, the antifungal susceptibility of planktonic cells, biofilm formation and susceptibility of biofilms to antifungal drugs and the secretion of key molecules, such as hydrolytic enzymes, hemolysins and siderophores, were evaluated by classical methodologies. Our results revealed that 7 (28%) isolates were molecularly identified as Ch, 7 (28%) as Chv and 11 (44%) as Cd. Sixteen (64%) fungal isolates were recovered from blood. Regarding the antifungal susceptibility test, the planktonic cells were resistant to (i) fluconazole (100% of Ch and Chv, and 72.7% of Cd isolates), itraconazole and voriconazole (85.7% of Ch and Chv, and 72.7% of Cd isolates); (ii) no breakpoints were defined for posaconazole, but high MICs were observed for 85.7% of Ch and Chv, and 72.7% of Cd isolates; (iii) all isolates were resistant to amphotericin B; and (iv) all isolates were susceptible to echinocandins (except for one isolate of Cd) and to flucytosine (except for two isolates of Cd). Biofilm is a well-known virulence and resistant structure in Candida species, including the C. haemulonii complex. Herein, we showed that all isolates were able to form viable biofilms over a polystyrene surface. Moreover, the mature biofilms formed by the C. haemulonii species complex presented a higher antifungal-resistant profile than their planktonic counterparts. Secreted molecules associated with virulence were also detected in our fungal collection: 100% of the isolates yielded aspartic proteases, hemolysins and siderophores as well as phospholipase (92%), esterase (80%), phytase (80%), and caseinase (76%) activities. Our results reinforce the multidrug resistance profile of the C. haemulonii species complex, including Brazilian clinical isolates, as well as their ability to produce important virulence attributes such as biofilms and different classes of hydrolytic enzymes, hemolysins and siderophores, which typically present a strain-dependent profile.

Development of DNA Bio-chip for Detection of Mutations of rpoB, embB and inhA Genes inDrug-Resistant Mycobacterium Tuberculosis.

Drug-resistant (DR) tuberculosis (TB) is a global threat to health security and TB control programs. Since conventional drug susceptibility testing (DST) takes several weeks, we have developed a molecular method for the rapid identification of DR strains of Mycobacterium Tuberculosis (M.tb) utilizing DNA bio-chips. DNA bio-chips were prepared by immobilizing oligonucleotides (probes) on highly microporous polycarbonate track-etched membranes (PC-TEM) as novel support. Bio-chip was designed to contain 15 specific probes to detect mutations in three genes (rpoB, embB, and inhA). A sensitive and specific chemiluminescence based bio-chip assay was developed based on multiplex PCR followed by hybridization on bio-chip. Fifty culture isolates were used to evaluate the ability of in-house developed bio-chip to detect the mutations. Bio-chip analysis shows that 37.7% of samples show wild type sequences, 53.3% of samples were monoresistance showing resistance to either rifampicin (RMP), isoniazid (INH), or ethambutol (EMB). 4.4% of samples were polydrug resistant showing mutations in both the rpoB gene and embB gene while 4.4% of samples were multidrug-resistant (MDR), harboring mutations in the rpoB and inhA genes. The results were compared with DST and sequencing. Compared to sequencing, bio-chip assay shows a sensitivity of 96.5% and specificity of 100% for RMP resistance. For EMB and INH, the results were in complete agreement with sequencing. This study demonstrates the first-time use of PC-TEMs for developing DNA bio-chip for the detection of mutations associated with drug resistance in M.tb. Developed DNA bio-chip accurately detected different mutations present in culture isolates and thus provides detailed and reliable data for clinical diagnosis.

Metagenomics of Cariogenic Microbiota in Primary Teeth with Deep Dentinal Caries: An Illumina-Based Sequencing Study

Discrepancy in the dynamic metabolic progression of the dental biofilm leads to dental caries, which is a polymicrobial infection. The homeostasis of the biofilm is unaffected and is not apparent unless there is an imbalance in the mineralization pattern of the supragingival biofilm. Literature provides numerous facts on microorganisms related to dental caries. Pathologic colonizers in the early stages of tooth decay involve organisms like Streptococcus oralis, subsequent adherence of Streptococcus sobrinus, and Streptococcus mutans[1]. Reports suggest the association of highly complex, variable, and uncultivable microbial population with dental caries, even though prime organisms are commonly known to be allied with disease succession. Association of mutans group streptococci and lactobacilli with the initiation of caries and early caries development have been well documented [2]. Studies have shown transition of early caries causing gram-positive primary facultative bacteria to anaerobic gram-positive rods and cocci and gram-negative rods as the lesion progresses to deep carious lesion[3]. Mapping of bacterial taxa revealed up to 14 phyla along with 101 genera. This comprised around 706 diverse OTUs (Operational Taxonomic Units). Caries progression at various stages and bacterial profile of different species associated with several disease have been observed. Some of the observed species found abundantly in whitespot lesion and dentin lesion of both dentitions were S. Parasanguinis and S. salivarius[4]. Molecular methods for bacterial identification and enumeration are performed routinely to precisely study bacterial species associated with dental caries, including those that are not presently cultivable.

Simple and rapid method for molecular identification of Konosirus punctatus and Clupanodon thrissa

Simple and rapid method for molecular identification of Konosirus punctatus and Clupanodon thrissa

Establishment of male‐specific sequence‐tagged site markers in Asparagus officinalis: An efficient tool for sex identification

AbstractGarden asparagus is a major dioecious plant that serves as a model for understanding sex determination and sex chromosome evolution. The shared cytogenetic homology of the sex chromosomes, with the exception of a mega base region missing from the X chromosome, has considerably hindered the identification of male and female plants. In this study, two male‐specific sequence‐tagged site (STS) markers Asp‐YS1 and Asp‐YS2, located on the Y chromosome, were discovered for early sex identification. The Asp‐YS1 and Asp‐YS2 markers were particularly effective in amplifying a fragment of 490‐bp and 296‐bp, respectively. Most of the male progenies of selected cultivars, and one homozygous male cultivar (‘Y14’), showed male‐specific amplification using these two markers. Of the 24 tested cultivars, the results of 21 cultivars demonstrated male‐specific amplification using the newly identified STS markers suggesting greater applicability and higher specificity over the previously existed Asp1‐T7sp marker, which is limited to some commercial cultivars. These findings presented the establishment of an efficient molecular method for sex identification in asparagus, enabling the rapid selection of male parental lines in future breeding programmes.

Detection of bacterial agents causing prostate infection by culture and molecular methods from biopsy specimens.

Background and Objectives:Prostatitis affects about 16% of men in their lifetime and sometimes leading to prostate cancer. Bacterial infections are the most common causes of prostatitis. Diagnosis of the causative agents of bacterial prostate infections plays an essential role in timely treating and preventing secondary complications. This study isolated bacterial infectious agents in patients’ surgical prostate and evaluated them by routine and molecular microbiological methods.Materials and Methods:In this cross-sectional study, 72 prostate biopsy specimens were collected from the Orology Departmen of hospitals of Qazvin University of Medical Sciences. All samples were cultured in aerobic and anaerobic conditions. Antibiotic susceptibility test by Kirby-Bauer standard method was performed for all isolated bacteria. In addition, all isolated bacteria were identified using 16S rDNA PCR and sanger sequencing methods. Also, TaqMan real-time PCR was applied to detect Ureaplasm aurealyticum, Mycoplasma hominins, and Mycoplasma genitalium.Results:In conventional culture method, out of 18 positive samples, 15 samples (83.3%) were Gram-negative bacteria and 3 samples (16.6%) were Gram-positive bacteria, containing Escherichia coli (55.5%), Klebsiella pneumoniae (11.1%), Enterobacter cloacae (5.5%), Pseudomonas aeruginosa (11.1%), Staphylococcus aureus (11.1%), and Enterococcus faecalis (5.5%). The results of molecular identification methods were the same as conventional culture results. Also, four patients were Ureaplasm aurealyticum, and three patients were positive for Mycoplasma hominis.Conclusion:Most bacteria isolated from prostate specimens belonged to the Enterobacteriaceae family, especially Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae. Staphylococcus aureus and Enterococcus faecalis were cocci isolated in the specimens too. Also, Ureaplasma urealyticum, and Mycoplasma hominis were identified in prostatitis.

Design and optimization of a 16S microbial qPCR multiplex for the presumptive identification of feces, saliva, vaginal and menstrual secretions.

Molecular methods for body fluid identification have been extensively researched in the forensic community over the last decade, mostly focusing on RNA‐based methods. Microbial DNA analysis has long been used for forensic applications, such as postmortem interval estimations, but only recently has it been applied to body fluid identification. High‐throughput sequencing of the 16S ribosomal RNA gene by previous research groups revealed that microbial signatures and abundances vary across human body fluids at the genus and/or species taxonomic level. Since quantitative PCR is still the current technique used in forensic DNA analysis, the purpose of this study was to design a qPCR multiplex targeting the 16S gene of Bacteroides uniformis, Streptococcus salivarius, and Lactobacillus crispatus that can distinguish between feces, saliva, and vaginal/menstrual secretions, respectively. Primers and probes were designed at the species level because these bacteria are highly abundant within their respective fluid. The validated 16S triplex was evaluated in DNA extracts from thirty donors of each body fluid. A classification regression tree model resulted in 96.5% classification accuracy of the population data, which demonstrates the ability of this 16S triplex to presumptively identify these fluids with high confidence at the quantification step of the forensic workflow using minimal input volume of DNA extracted from evidentiary samples.

Development of a Rapid and Efficient RPA-CRISPR/Cas12a Assay for Mycoplasma pneumoniae Detection.

Mycoplasma pneumoniae (MP) is a one of most common pathogen in causing respiratory infection in children and adolescents. Rapid and efficient diagnostic methods are crucial for control and treatment of MP infections. Herein, we present an operationally simple, rapid and efficient molecular method for MP identification, which eliminates expensive instruments and specialized personnel. The method combines recombinase polymerase amplification (RPA) with clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated proteins (Cas) 12a-based detection, with an optimal procedure less than 1 h from sample to result including DNA extraction (25 min), RPA reaction (39°C for 15-20 min), CRISPR/Cas12a detection (37°C for 10 min) and visual detection by naked eyes (2 min). This diagnostic method shows high sensitivity (two copies per reaction) and no cross-reactivity against other common pathogenic bacteria. Preliminary evaluation using 201 clinical samples shows sensitivity of 99.1% (107/108), specificity of 100% (93/93) and consistency of 99.5% (200/201), compared with real-time PCR method. The above data demonstrate that our developed method is reliable for rapid diagnosis of MP. In conclusion, the RPA-CRISPR/Cas12a has a great potential to be as a useful tool for reliable and quick diagnosis of MP infection, especially in primary hospitals with limited conditions.

SSR Loci Analysis in Transcriptome and Molecular Marker Development in Polygonatum sibiricum.

To study the SSR loci information and develop molecular markers, a total of 435,858 unigenes in transcriptome of Polygonatum sibiricum were used to explore SSR. The distribution frequency of SSR and the basic characteristics of repeat motifs were analyzed using MISA software, and SSR primers were designed by Primer 3.0 software and then validated by PCR. Moreover, the gene function analysis of SSR Unigene was obtained by Blast. The results showed that 112,728 SSR loci were found in the transcriptome of Polygonatum sibiricum, which distributed in 435,858 unigenes with a distribution frequency of 25.86%. Mo-nucleotide and Di-nucleotide repeat were the main types, accounted for 83.83% of all SSRs. The repeat motifs of A/T and AC/GT were the predominant repeat types of Mo-nucleotide and Di-nucleotide, respectively. A total of 113,305 pairs of SSR primers with the potential to produce polymorphism were designed for maker development. One hundred and fifty-four of the 500 randomly selected primers not only produced fragments with expected molecular size but also had high polymorphism, which could accurately separate the tested varieties. The gene function of unigenes containing SSR was mostly related to the molecular function of Polygonatum sibiricum. The SSR markers in transcriptome of Polygonatum sibiricum show rich type, strong specificity, and high potential of polymorphism, which will benefit the candidate gene mining and marker-assisted breeding. The developed markers can also provide technical methods for molecular identification of intraspecific species of Polygonatum Mill. and maker-assisted breeding of superior varieties of Polygonatum Mill.

Compatibility of Diatom Valve Records With Sedimentary Ancient DNA Amplicon Data: A Case Study in a Brackish, Alkaline Tibetan Lake

Lake sediments represent valuable and widely used archives for tracking environmental and biotic changes over time. Past aquatic communities are traditionally studied via morphological identification of the remains of organisms. However, molecular identification tools, such as DNA metabarcoding, have revolutionized the field of biomonitoring by enabling high-throughput and fast identification of organisms from environmental samples (e.g., sediments and soil). Sedimentary ancient DNA (sedaDNA) metabarcoding, an approach to track the biodiversity of target organisms from sediment cores, spanning thousands of years, has been successfully applied in many studies. However, researchers seldom explore how well the signals from sedaDNA data correlate with the fossil records of target organisms. This information is essential to infer past environmental conditions and community changes of bioindicators when the increasingly popular molecular identification method, metabarcoding, is desired instead of a morphological identification approach. In this study, we explore the correlations of diatom valve records across the last ∼940 years with the diatom sedaDNA metabarcoding data from the same sediment core from lake Nam Co (Tibetan Plateau). Overall, the results from valve vs. sedaDNA data revealed concordant diatom richness as well as community patterns. However, several mismatches in the diatom taxonomic composition existed between the data sets. In general, sedaDNA data harbored much higher diatom diversity, but due to the lack of reference sequences in public databases, many molecular units (amplicon sequence variants) remained unclassified to lower taxonomic levels. As our study lake, Nam Co, is characterized by brackish water and alkaline pH, some likely cases for the observed taxonomic composition mismatches may be due to a valve dissolution issue. Nevertheless, significant drivers for the diatom richness and community structure largely corresponded between data sets. Both valve and sedaDNA data demonstrated similar breakpoints for historical diatom community shifts. A particularly strong shift in the diatom community structure occurred after ∼1950 CE, which may be associated with abrupt environmental changes on the Tibetan Plateau. Altogether, our study indicates that environmentally driven signals reflected by the diatom communities are successfully recovered via microfossil as well as molecular identification methods.

Dispersal of Aphanoascus keratinophilus by the rook Corvus frugilegus during breeding in East Poland

The process of dispersal of the potentially disease-causing, geophilic and keratinolytic fungal strain Aphanoascus keratinophilus (the perfect, sexual stage of Chrysosporium keratinophilum) by the rook Corvus frugilegus was studied. The source of A. keratinophilus strains was pellets of the rook, thus far not considered a carrier of this particular opportunistic pathogen. Pellets collected from breeding colonies of rooks were analysed in terms of the occurrence of keratinolytic fungi with the application of the native keratin bait method. Among the 83 rook pellets analysed, 24 (29%) were infected by keratinophilic fungi. Pure cultures of the fungi were identified to species based on traditional morphological features. Traditional mycological identification was verified by the PCR–RFLP molecular identification method as well as DNA sequencing. The obtained results showed the presence of 90 Aphanoascus keratinophilus strains, 6 Chrysosporium tropicum strains, and 3 Chrysosporium pannicola strains. The PCR melting profile (PCR-MP) method was used to identify intraspecies variations of the 90 analysed A. keratinophilus strains. The dispersal of genotypes and possible pathways of A. keratinophilus dispersal and infection via rook pellets were analysed.

Combining of molecular 16S rRNA gene and metabolic fingerprinting through biolog system for the identification of streptomycetes in Saudi Arabia

Many of the conventional identification methods for the group of streptomycetes deal with attempts to reveal their metabolic activity as well as their genetic makeup. The current study dealt with the possibility of combining one of the most prevalent molecular identification methods of bacteria, which is 16S rRNA gene sequencing along with the use of some basic standard methods and linking them with the Biolog system for bacterial identification. The study was conducted on the isolated Streptomyces (isolate EA3) with an ability of cellulose utilization, a well-known activity of the genus Streptomyces in nature. Identification of the isolate was achieved by standard phenotypic methods. The isolate was metabolically fingerprinted using the Biolog MicroPlate. Molecular identification was also performed using analysis of 16 s rRNA. The characteristic band of about 1487 bp was obtained. The sequencing analysis was performed and revealed that the isolate is very close to Streptomyces cinereoruber. The obtained sequence, 405 bp, was deposited in GenBank under accession, MT907291.1. The study concludes on the importance of linking the molecular and metabolic fingerprints of this genus of bacteria because of its enormous role in the natural processes and its numerous benefits.

POS-550 MASS-SPECTROMETRIC IDENTIFICATION OF POST-TRANSLATIONAL GUANIDINYLATED YB-1 PROTEIN IN PATIENTS WITH SYSTEMIC LUPUS ERYTHEMATOSUS

With continuous identification of post-translational modified isoforms of proteins, it is becoming increasingly clear that post-translational modifications limit or modify the biological functions of native proteins are majorly involved in development of various chronic disease. This is mostly due to technically advanced molecular identification and quantification methods, mainly based on mass spectrometry. Mass spectrometry has become one of the most powerful tools for the identification of lipids.

Genetic diversity and phylogenetic analysis of Khiew-Phalee chickens based on mitochondrial DNA cytochrome b gene sequences

Abstract. Phromnoi S, Lertwatcharasarakul P, Phattanakunanan S. 2021. Genetic diversity and phylogenetic analysis of Khiew-Phalee chickens based on mitochondrial DNA cytochrome b gene sequences. Biodiversitas 23: 750-756. Khiew-Phalee Chickens (Gallus gallus) is a Thai indigenous breed, locally recognized for its economic value as an important source of revenue generation. As the Khiew-Phalee chicken cannot be accurately identified by morphological characterization, molecular methods for species identification are generally thought to be complementary and more effective than relying on the method alone. Molecular characterization of Khiew-Phalee Chickens was investigated using mitochondrial DNA (mtDNA) cytochrome b (Cyt b) gene analysis. Partial sequences of the 1,037 bp Cyt b gene were determined on 55 specimens of Khiew-Phalee chickens. The breed's genetic diversity was assessed based on the number of polymorphic sites, number of haplotypes, haplotype diversity, nucleotide diversity, and average number of differences. In addition, a neighbor-joining (NJ) haplotype tree was constructed based on Kimura's two-parameter model. Haplotype and nucleotide diversity of Khiew-Phalee chickens were 0.597 and 0.00068, respectively. Five haplotypes were identified from 4 polymorphic sites with polymorphism at nucleotides 501, 507, 543, and 1,095. Phylogenetic analysis with the inclusion of the Cyt b gene nucleotide sequences from Khiew-Phalee chickens and native chickens from the GenBank database showed that Khiew-Phalee chickens belonged to Gallus gallus. It was closest to the indigenous chicken in South East Asia. The phylogenetic tree indicated the genetic variability of the investigated Khiew-Phalee chicken populations. This study may help future researchers and livestock breeders to design a breeding program based on a better understanding of the genetic diversity and history of indigenous breeds.

An incubation water eDNA method for a non-destructive rapid molecular identification of Pinna nobilis and Pinna rudis bivalve juveniles

The pen shell Pinna nobilis is critically endangered due to a disease that has affected all open water populations since late 2016. Collection of early spats is considered a fundamental step for pen shell conservation. However, the identification between P. nobilis and P. rudis juveniles by morphology is a very difficult task. Furthermore, due to the small size of juveniles and high sensitivity to handling, the sampling for this purpose must not damage individuals. As a consequence, the application of molecular techniques for conservation strategies to identify threatened and endangered bivalve species is every day more and more necessary.In this study, we present the development of a multiplex-PCR procedure for the rapid identification of two Pinna species from eDNA water samples. Using species-specific primers, designed in the rRNA16S and rRNA12S mitochondrial genes, identification of species was obtained by cellular or extracellular DNA dissolved in water and differentiated based on the size of the amplified DNA fragments.• Development of a molecular multiplex-PCR procedure for the rapid identification of two Pinna species from eDNA water samples• Using specie-specific primers, the different species can be differentiated basing on the size of the amplified DNA fragments• This technique removes many of the limitations commonly associated with sampling of threatened and endangered juvenile bivalves for conservation strategies (sampling does not damage individuals).

Эпидемиологическое значение хронической инфекции легких, вызванной бактериями рода Achromobacter, у пациентов с муковисцидозом

Achromobacter is a genus of nonfermenting gram-negative bacteria that have particular clinical importance for patients with cystic fibrosis. Objective. To analyze epidemiological role of chronic lung infection caused by Achromobacter in patients with cystic fibrosis in Russia. Patients and methods. We examined 142 Achromobacter spp. isolates from sputum samples and throat swabs collected from 89 children and 53 adults. We used bacteriological and molecular methods (PCR, WGS) for identification and typing. Results. Our results demonstrated that newly identified chronic lung infection caused by Achromobacter spp. can be both nosocomial and community-acquired. Pathogens with different genotypes (ST36, ST261, ST184) are transmissive. Types ST36 and ST261 are found in several continents. Since 33.3% of patients infected with Achromobacter spp. were also found to have A. ruhlandii ST36, it can be defined as an epidemic clone. Our findings suggest high clinical and epidemiological importance of Achromobacter spp. Key words: Achromobacter spp., cystic fibrosis, a chronic lung infection

Emerging and reemerging forms of Trypanosoma cruzi transmission.

This review aims to update and discuss the main challenges in controlling emergent and reemergent forms of Trypanosoma cruzi transmission through organ transplantation, blood products and vertical transmission in endemic and non-endemic areas as well as emergent forms of transmission in endemic countries through contaminated food, currently representing the major cause of acute illness in several countries. As a neglected tropical disease potentially controllable with a major impact on morbimortality and socioeconomic aspects, Chagas disease (CD) was approved at the WHO global plan to interrupt four transmission routes by 2030 (vector/blood transfusion/organ transplant/congenital). Implementation of universal or target screening for CD are highly recommended in blood banks of non-endemic regions; in organ transplants donors in endemic/non-endemic areas as well as in women at risk from endemic areas (reproductive age women/pregnant women-respective babies). Moreover, main challenges for surveillance are the application of molecular methods for identification of infected babies, donor transmitted infection and of live parasites in the food. In addition, the systematic recording of acute/non-acute cases and transmission sources is crucial to establish databases for control and surveillance purposes. Remarkably, antiparasitic treatment of infected reproductive age women and infected babies is essential for the elimination of congenital CD by 2030.

Molecular identification guided process design for deep removal of fluoride from electroplating effluent

This study developed a molecular identification method for fluoride in industrial wastewater and explored the adsorptive removal of fluoroborate by strongly basic ion exchange resin.