Genotypic Testing Research Articles

HIV Drug Resistance Patterns and Characteristics Associated with Clinically Significant Drug Resistance among Children with Virologic Failure on Antiretroviral Treatment in Kenya: Findings from the Opt4Kids Randomized Controlled Trial.

Increasing HIV drug resistance (DR) among children with HIV (CHIV) on antiretroviral treatment (ART) is concerning. CHIV ages 1-14 years enrolled from March 2019 to December 2020 from five facilities in Kisumu County, Kenya, were included. Children were randomized 1:1 to control (standard-of-care) or intervention (point-of-care viral load (POC VL) testing every three months with targeted genotypic drug resistance testing (DRT) for virologic failure (VF) (≥1000 copies/mL)). A multidisciplinary committee reviewed CHIV with DRT results and offered treatment recommendations. We describe DR mutations and present logistic regression models to identify factors associated with clinically significant DR. We enrolled 704 children in the study; the median age was 9 years (interquartile range (IQR) 7, 12), 344 (49%) were female, and the median time on ART was 5 years (IQR 3, 8). During the study period, 106 (15%) children had DRT results (84 intervention and 22 control). DRT detected mutations associated with DR in all participants tested, with 93 (88%) having major mutations, including 51 (54%) with dual-class resistance. A history of VF in the prior 2 years (adjusted odds ratio (aOR) 11.1; 95% confidence interval (CI) 6.3, 20.0) and less than 2 years on ART at enrollment (aOR 2.2; 95% CI 1.1, 4.4) were associated with increased odds of major DR. DR is highly prevalent among CHIV on ART with VF in Kenya. Factors associated with drug resistance may be used to determine which children should be prioritized for DRT.

Influence of pharmacogenetics on the diversity of response to statins associated with adverse drug reactions.

Statins are one of the most prescribed medications in developed countries as the treatment of choice for reducing cholesterol and preventing cardiovascular diseases. However, a large proportion of patients experience adverse drug reactions, especially myotoxicity. Among the factors that influence the diversity of response, pharmacogenetics emerges as a relevant factor of influence in inter-individual differences in response to statins and can be useful in the prevention of adverse drug effects. A systematic review was performed of current knowledge of the influence of pharmacogenetics on the occurrence and prevention of statin-associated adverse reactions and clinical benefits of preemptive pharmacogenetics testing. Genetic variants SLCO1B1 (rs4149056) for all statins; ABCG2 (rs2231142) for rosuvastatin; or CYP2C9 (rs1799853 and rs1057910) for fluvastatin are associated with an increase in muscle-related adverse effects and poor treatment adherence. Besides, various inhibitors of these transporters and biotransformation enzymes increase the systemic exposure of statins, thereby favoring the occurrence of adverse drug reactions. The clinical preemptive testing of this pharmacogenetic panel would largely prevent the incidence of adverse drug reactions. Standardized methods should be used for the identification of adverse effects and the performance and interpretation of genotyping test results. Standardization would allow to obtain more conclusive results about the association between SLCO1B1, ABCG and CYP2C9 variants and the occurrence of adverse drug reactions. As a result, more personalized recommendations could be established for each statin.

HIV-1 drug resistance in people on dolutegravir-based antiretroviral therapy: a collaborative cohort analysis

The widespread use of the integrase strand transfer inhibitor (INSTI) dolutegravir in first-line and second-line antiretroviral therapy (ART) might facilitate emerging resistance. The DTG RESIST study combined data from HIV cohorts to examine patterns of drug resistance mutations (DRMs) and identify risk factors for dolutegravir resistance. We included cohorts with INSTI resistance data from two collaborations (ART Cohort Collaboration, International epidemiology Databases to Evaluate AIDS in Southern Africa), and the UK Collaborative HIV Cohort. Eight cohorts from Canada, France, Germany, Italy, the Netherlands, Switzerland, South Africa, and the UK contributed data on individuals who were viraemic on dolutegravir-based ART and underwent genotypic resistance testing. Individuals with unknown dolutegravir initiation date were excluded. Resistance levels were categorised using the Stanford algorithm. We identified risk factors for resistance using mixed-effects ordinal logistic regression models. We included 599 people with genotypic resistance testing on dolutegravir-based ART between May 22, 2013, and Dec 20, 2021. Most had HIV-1 subtype B (n=351, 59%), a third had been exposed to first-generation INSTIs (n=193, 32%), 70 (12%) were on dolutegravir dual therapy, and 18 (3%) were on dolutegravir monotherapy. INSTI DRMs were detected in 86 (14%) individuals; 20 (3%) had more than one mutation. Most (n=563, 94%) were susceptible to dolutegravir, seven (1%) had potential low, six (1%) low, 17 (3%) intermediate, and six (1%) high-level dolutegravir resistance. The risk of dolutegravir resistance was higher on dolutegravir monotherapy (adjusted odds ratio [aOR] 34·1, 95% CI 9·93-117) and dolutegravir plus lamivudine dual therapy (aOR 9·21, 2·20-38·6) compared with combination ART, and in the presence of potential low or low (aOR 5·23, 1·32-20·7) or intermediate or high-level (aOR 13·4, 4·55-39·7) nucleoside reverse transcriptase inhibitor (NRTI) resistance. Among people with viraemia on dolutegravir-based ART, INSTI DRMs and dolutegravir resistance were rare. NRTI resistance substantially increased the risk of dolutegravir resistance, which is of concern, notably in resource-limited settings. Monitoring is important to prevent resistance at the individual and population level and ensure the long-term sustainability of ART. US National Institutes of Health, Swiss National Science Foundation.

Detection of carbapenemase production in gram negative bacilli in medical and surgical intensive care unit patients in a tertiary care hospital

Background: Carbapenem-resistant Enterobacterales (CRE) are an urgent global public health problem. CRE infections are associated with high mortality and have limited available effective treatment. Carbapenemase enzymes are encoded by genes on mobile genetic elements, such as plasmids, which are highly transmissible between organisms and increase the potential spread of resistance. The study aim was to detect carbapenemase production in Gram negative bacilli by phenotypic (CarbaNP and mCIM only or in conjunction with eCIM) and genotypic (Xpert Carba-R) methods. Material and methods: The various clinical samples were processed as per standard recommended procedures. Identification and antibiotic susceptibility test were done by using GN cards and AST N280 & AST N281 cards of Vitek 2 Compact (bioMérieux) respectively. Phenotypic (CarbaNP and mCIM only or in conjunction with eCIM) and genotypic (Xpert Carba-R) methods were used for detection of carbapenemases. Result: 144 carbapenem-resistant Gram-negative bacilli isolated from various ICUs includes Klebsiella pneumoniae 52.7% (76/144) followed by Escherichia coli 18.05% (26/144) and 15.9% (23/144) Acinetobacter baumannii. Phenotypic test (CarbaNP and mCIM and or in conjunction with eCIM) showed sensitivity of 90%, 100% and 93.75% respectively. Genotypic test of 40 isolates showed predominant expression of NDM in 82.5% (33/40) isolates followed by OXA-48 in 40% (16/40). Conclusion: The study showed mCIM as the most useful diagnostic test with less economic burden to the patients. There is an urgent need for more sensitive, rapid, highly precise and accurate genotypic test which is less expensive and less labor-intensive. Keywords: carbapenem resistant; Enterobacterales; mCIM; eCIM; Xpert Carba-R

Resistance testing of several genotypes of Kamba Local Paddy (Oryzae sativa L.) to leaf blight caused by Xanthomonas oryzae pv. oryzae

One of the important diseases of paddy plants is bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae (Xoo). Kamba is one of the local paddy plants in Central Sulawesi’s germplasm collection that is superior in origin from the Lore (Bada, Napu and Lindu). Until now, no scientific testing has been conducted on the resistance of local kamba paddy to leaf blight. This study aims to compare the level of resistance of several kamba paddy genotypes to leaf blight caused by the bacterium Xoo This research was conducted at the Disease Laboratory and Plant Pests and Diseases Gauze House, Faculty of Agriculture, Tadulako University, Palu. Stages of research implementation include propagation of Xoo, Preparation of test plants, and Transmission of Xoo. Observation variables include symptoms on plants, incubation period, disease severity and level of resistance to Xoo. The results showed that Paddy cultivars Kamba Bulili Kasar and Kamba Bulili Halus showed resistance reactions to Bacterial Leaf Blight (HDB) caused by Xoo bacteria. Both paddy cultivars also showed a disease incubation period of 6 days, longer than the susceptible comparison varieties.

Association of Single Nucleotide Polymorphism in VDR, GC Globulin and CYP2R1 with the Risk of Esophageal Cancer.

The proactive role of vitamin D has been well determined in different cancers. The protein that encodes the components of the vitamin D metabolism could appear to play a pivotal role in vitamin D stability and its maintenance. A polymorphism in vitamin-D-receptor (VDR), carrier globulin/binding protein (GC) and cytochrome P-450 family 2, subfamily R, polypeptide 1 (CYP2R1) genes has been predicted to be associated with the development of cancer. This study was designed to detect the association of VDR, GC Globulin and CYP2R1 gene polymorphism with the risk of esophageal cancer in the North-east Indian population. To carry out the study, a total of 100 patients diagnosed with esophageal cancer and 101 healthy controls were enrolled. In a case-control manner, all samples were subjected to do genotype testing for known SNPs on the VDR (rs1544410), GC (rs4588), and CYP2R1 (rs10741657) genes using Restriction-fragment length polymorphism (RFLP) followed by Sanger sequencing. The collected demographic and clinical data were analysed using the statistical software package SPSS v22.0. The VDR haplotype heterozygous TC was found strongly associated with the carcinoma group (OR:1.09, 95%CI:0.67-1.75). The risk factors analysis using the GC globulin rs4588 phenotype, found a positive correlation in terms of mutant AA's harmful influence on the cancer cohort (OR = 1.125, OR=1.125, 95% CI, 0.573-2.206). The influence of the CYP2R1 rs10741657 polymorphism on the malignant cohort revealed that the GG mutant had a significant negative influence on the carcinoma, has an influential role in disease severity ( OR:1.736, at 95% CI; 0.368-8.180). In conclusion, this study revealed the potential association of VDR gene polymorphism in the progression and development of esophageal cancer in north east Indian population cohort.

Computer aided diagnosis of neurodevelopmental disorders and genetic syndromes based on facial images – A systematic literature review

Neurodevelopment disorders can result in facial dysmorphisms. Therefore, the analysis of facial images using image processing and machine learning techniques can help construct systems for diagnosing genetic syndromes and neurodevelopmental disorders. The systems offer faster and cost-effective alternatives for genotyping tests, particularly when dealing with large-scale applications. However, there are still challenges to overcome to ensure the accuracy and reliability of computer-aided diagnosis systems. This article presents a systematic review of such initiatives, including 55 articles. The main aspects used to develop these diagnostic systems were discussed, namely datasets - availability, type of image, size, ethnicities and syndromes - types of facial features, techniques used for normalization, dimensionality reduction and classification, deep learning, as well as a discussion related to the main gaps, challenges and opportunities.

Exploring programmatic indicators of tuberculosis control that incorporate routine Mycobacterium tuberculosis sequencing in low incidence settings: a comprehensive (2017–2021) patient cohort analysis

Routine whole genome sequencing of Mycobacterium tuberculosis has been implemented with increasing frequency. However, its value for tuberculosis (TB) control programs beyond individual case management and enhanced drug resistance detection has not yet been explored. We analysed routine sequencing data of culture-confirmed TB cases notified between 1st January 2017 and 31st December 2021 in New South Wales (NSW), Australia. Genomic surveillance included evidence of local TB transmission, defined by single nucleotide polymorphism (SNP) clustering over a variable (0-25) SNP threshold, and drug resistance conferring mutations. M.tuberculosis sequences from 1831 patients were examined, representing 64.8% of all notified TB cases and 96.2% of culture-confirmed cases. Applying a traditional 5-SNP cluster threshold identified 62 transmission clusters with 183 clustered cases; 101/183 (55.2%) had 0 SNP differences. Cluster assessment over a 5-year period, using a 5-SNP threshold, provided a comprehensive overview of likely recent transmission within NSW, Australia, as an indicator of local TB control. Genotypic drug susceptibility testing (DST) was highly concordant with phenotypic DST and provided a 6.8% increase in antimycobacterial resistance detection. Importantly, it detected mutations missed by routine molecular tests. Lineage 2 strains were more likely to be drug resistant (p<0.0001) and locally transmitted if drug resistant (p<0.0001). Performing routine prospective WGS in a low incidence country like Australia, provides genomically informed programmatic indicators of local TB control. A rolling 5-year cluster assessment reflects epidemic containment and progress towards 'zero TB transmission'. Genomic DST also provides valuable information for clinical care and drug resistance surveillance. NHMRC Centre for Research Excellence in Tuberculosis (www.tbcre.org.au) and NSW Health Prevention Research Support Program.

Recommendations on data sharing in HIV drug resistance research.

• Human immunodeficiency virus (HIV) drug resistance has implications for antiretroviral treatment strategies and for containing the HIV pandemic because the development of HIV drug resistance leads to the requirement for antiretroviral drugs that may be less effective, less well-tolerated, and more expensive than those used in first-line regimens. • HIV drug resistance studies are designed to determine which HIV mutations are selected by antiretroviral drugs and, in turn, how these mutations affect antiretroviral drug susceptibility and response to future antiretroviral treatment regimens. • Such studies collectively form a vital knowledge base essential for monitoring global HIV drug resistance trends, interpreting HIV genotypic tests, and updating HIV treatment guidelines. • Although HIV drug resistance data are collected in many studies, such data are often not publicly shared, prompting the need to recommend best practices to encourage and standardize HIV drug resistance data sharing. • In contrast to other viruses, sharing HIV sequences from phylogenetic studies of transmission dynamics requires additional precautions as HIV transmission is criminalized in many countries and regions. • Our recommendations are designed to ensure that the data that contribute to HIV drug resistance knowledge will be available without undue hardship to those publishing HIV drug resistance studies and without risk to people living with HIV.

Phenotype and genotype analysis for Helicobacter pylori antibiotic resistance in outpatients: a retrospective study.

To investigate the antibiotic resistance of Helicobacter pylori (H. pylori) in outpatients and to explore the consistency between genotype and phenotype of H. pylori antibiotic resistance. A retrospective study on outpatients screened with urea breath test for H. pylori infection in Nanjing First Hospital from April 2018 to January 2022. Patients who tested positive underwent a consented upper endoscopy, and the H. pylori infection was confirmed by rapid urease test (RUT) and H. pylori culture. For antibiotic resistance phenotype analysis, the H. pylori strains isolated from gastric biopsy were tested for antibiotic resistance phenotype by the Kirby-Bauer disk diffusion test. In addition, the antibiotic resistance genotype of isolated H. pylori was tested with a real-time polymerase chain reaction. A total of 4,399 patients underwent H. pylori infection screening, and 3,306 H. pylori strains were isolated. The antibiotic resistance phenotype test revealed that the resistance rates of metronidazole (MTZ), clarithromycin (CLR), levofloxacin (LEV), amoxicillin (AMX), furazolidone (FR), and tetracycline (TE) were 74.58%, 48.61%, 34.83%, 0.76%, 0.27%, and 0.09%, respectively. Additionally, the antibiotic resistance genotype test revealed that rdxA gene mutation A610G (92.96%), A91G (92.95%), C92A (93.00%), and G392A (95.07%) were predominant in H. pylori with MTZ resistance; 23S rRNA gene mutation A2143G (86.47%) occurred in most H. pylori with CLR resistance; and gyrA gene mutation 87Ile/Lys/Tyr/Arg (97.32%) and 91Asn/Gly/Tyr (90.61%) were the most popular mutations in strains with LEV resistance. The phenotypic resistance and genotypic resistance to CLR (kappa value = 0.824) and LEV (kappa value = 0.895) were in good agreement. The history of eradication with MTZ, CLR, LEV, and AMX was correlated with H. pylori resistance. In short, this study demonstrated that drug resistance of H. pylori was mainly to MTZ, CLR, and LEV in local outpatients. Three drugs can be selected for increased MICs (Minimum Inhibitory Concentration) via single chromosomal mutations. In addition, the genotype could be used to predict the phenotypic H. pylori resistance to CLR and LEV. IMPORTANCE Helicobacter pylori is a key bacterium that causes stomach diseases. There was a high prevalence of H. pylori in the Chinese population. We analyzed the resistance phenotype and genotype characteristics of H. pylori in 4,399 outpatients at the First Hospital of Nanjing, China. We found a higher resistance rate to metronidazole (MTZ) , clarithromycin (CLR), and levofloxacin (LEV), and the genotype could be used to predict the phenotypic H. pylori resistance to CLR and LEV. This study provides information on H. pylori infection and also provides guidance for clinical doctors' drug treatment.

Abstract PR10: A decade in the making: Screening ctDNA detection in HPV-associated HNSCC

Abstract Introduction HPV-associated head and neck squamous cell carcinomas (HPV+HNSCC) release circulating tumor DNA (ctDNA), including ctHPVDNA, into the blood, which is detectable at the time of diagnosis in nearly all patients with HPV+HNSCC using droplet digital PCR (ddPCR)-based approaches. We have previously demonstrated proof of principle that using these approaches, ctHPVDNA is detectable in a minority of patients up to 3 years prior to diagnosis, suggesting HPV+HNSCC may be reliably detected many years prior to clinical detection when using more sensitive approaches. Here, we applied a custom whole genome hybrid-capture-based next generation sequencing (NGS) assay, termed HPVseek, to plasma samples collected years prior to HPV+HNSCC diagnosis to test this hypothesis. Method Plasma samples from 28 patients collected 1-11 years prior to HPV+HNSCC diagnosis were obtained from the Mass General Brigham Biobank, along with 1:1 age and gender matched controls. Cell free DNA was extracted and run on HPVseek and our existing ddPCR assay. Pre-existing cut-points for a positive result were applied. In cases where tissue blocks were available from the time of diagnosis, DNA was extracted and underwent HPVseek for confirmatory molecular fingerprinting of the virus. Result 19 of 28 patients (68%) had detectable ctHPVDNA with lead times ranging from 1.0 to 7.5 years. Of the 11 samples obtained between 1-3 years before diagnosis, 11(100%) were ctHPVDNA positive. All controls were negative. HPV genotype distribution in positive samples was: 16 (n=16), 35 (1), 56 (1), and 59 (1) in blood with genotypes matching clinical genotype testing in 27/28 cases. 11/28 (39%) were positive on ddPCR, all of which were also positive on NGS. Molecular fingerprinting of viral genomes revealed unique viral genomes in each case and matching viral genomes within paired blood and tissue blocks, ruling out contamination and demonstrating the same inciting virus pre-diagnosis to diagnosis. Seven patients had diagnostic and post-diagnostic blood samples available as part of our ongoing prospective ctHPVDNA studies, allowing continuous tracking of cancer status along a time of up to 12 years, providing a global picture of HPV+HNSCC. Conclusion Here we demonstrate that HPV+HNSCC is detectable in the blood as ctDNA &amp;gt;seven years prior to diagnosis, and was detectable in 100% of cases up to three years prior to diagnosis, using an ultrasensitive custom ctDNA NGS assay. Findings were orthogonally validated using molecular fingerprinting in tissue and blood. This study has broad implications for the field of liquid biopsy cancer screening, demonstrating perhaps the longest lead times ever reported for ctDNA detection prior to cancer clinical diagnosis and suggesting HPV+HNSCC could be screened for, detected, and potentially treated years before clinical cancer detection. Citation Format: Shun Hirayama, Brian Y. Zhao, Saskia Naegele, Vasileios Efthymiou, Julia Mendel, Adam Fisch, Michael S. Lawrence, Viktor Adalsteinsson, Anthony J. Iafrate, Dipon Das, Daniel L. Faden. A decade in the making: Screening ctDNA detection in HPV-associated HNSCC [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Innovating through Basic, Clinical, and Translational Research; 2023 Jul 7-8; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2023;29(18_Suppl):Abstract nr PR10.

Responses of β-thalassemia and compound heterozygote of Sickle/βthalassemia of BCL11A Gene Polymorphism in Pakistani Patients.

Beta-thalassemia major (β-Thal) and compound heterozygote of Sickle β-thalassemia (S-β Thal) are hereditary autosomal recessive disorders resulting from mutations or deletion in β-globin gene cluster. Patients with increased HbF levels having polymorphism at BCL11A site loci have shown clinical significance. The present study aimed to assess the frequency of BCL11A gene polymorphism in a study population of β-Thal, S-β Thal & Controls using Sanger sequencing leading to plot the HbF response of polymorphism with reference to wild type. The sample size of the study is n=180, groups were divided in Controls, β-thal & S-β thal. One ml blood was drawn from patients and controls to extract DNA for PCR amplification and BCL11A locus genotyping using Sanger sequencing. This study was carried out at Dow Research Institute of Biotechnology and Biomedical Sciences, for one year from March 2021 to February 2022. The HbF response of three groups is hyperbolic with 83 for β-Thal, 16 for S-β Thal and close to zero for controls. The frequency of heterozygous variant GA of BCL11A gene polymorphism is 51%. The frequency of homozygous variant GG is 49%. Complete absence of wild type AA in patient group. The frequency of BCL11A polymorphism in control group was 43% (with male 18% and female 21%) showing wild type status of 57%. The patient groups of SCD and Beta thalassemia are devoid of wild type status. The wild type status of BCL11A is 57% even in control population. Higher level of HbF in B-thalassemia and SCD and B Thalassemia is a cost-effective screening marker before switching to an expensive genotyping testing.

Dual Antiplatelet Therapy in Minor Stroke/Transient Ischemic Attack - An Updated Network Meta-Analysis.

We previously analyzed five trials on ticagrelor/aspirin versus clopidogrel/aspirin in patients with minor stroke/ TIA in a network meta-analysis. We updated our search and identified 311 new citations with one study for inclusion: CHANCE2 enrolled patients with CYP2C19 loss-of-function alleles and randomized them to ticagrelor/aspirin or clopidogrel/aspirin. Pooling of CHANCE2 with the original studies could not be completed due to violation of NMA assumptions, due to significant inconsistency. This suggests patients with CYP2C19 loss-of-function alleles represent a subpopulation that is inherently different from the general stroke population in their antiplatelet response. Results from CHANCE-2 may not be generalizable without genotype testing.

Usefulness and difficulties with the thiopurine pharmacogenomic NUDT15 genotyping test: Analysis of real-world data in Japan

The usefulness of NUDT15 genotyping as a pharmacogenomic test for thiopurine has been established. The first such test developed to date, NUDT15 genotyping was approved for reimbursement in Japan in February 2019 for all indicated patients. We retrospectively examined claims data in Japan and confirmed that the proportion of patients who undergo genotyping before initiating a new thiopurine regimen has increased; furthermore, genotyping has improved the rate of treatment continuation and reduced on-treatment hospitalization. However, the genotyping rate before thiopurine induction was >50% for patients with inflammatory bowel disease and <20% for those with other immune-related diseases, indicating significant variation by disease field. Additionally, over 10% of tests were found to have been performed inappropriately, such as multiple genotyping of the same patient or testing more than 2 weeks after starting treatment. Although NUDT15 genotyping for patients requiring thiopurine treatment has been shown to improve thiopurine treatment continuation rate, measures are required to address the systematic issues identified in our analysis.

Incidence and Patterns of Drug Resistance in Patients with Spinal Tuberculosis: a Prospective, Single-Center Study from a Tuberculosis-Endemic Country.

There is paucity of data on incidence and pattern of drug resistance in spinal TB. This prospective observational study was conducted to document the incidence and drug-resistance pattern among primary and presumptive resistant cases. 59 consecutive cases diagnosed clinico-radiologically (imaging) were grouped into Group A (n = 51, primary cases) and Group B (n = 8, presumptive resistant cases) based on pre-defined criteria (INDEX-TB guidelines). Tissue samples obtained percutaneously (37.29%, 22/59) and on surgery (62.71%, 37/59) were subjected to genotypic DST (CBNAAT, LPA) and phenotypic DST (BACTEC MGIT 960 culture and sensitivity using fixed critical concentration of drugs). Etiological diagnosis was ascertained in all. 13/51 (25.49%) in Group A, while 3/8 (37.5%) in Group B and 16/59 (27.12%) overall demonstrated drug resistance. 12/16 (75%) had no prior history of ATT intake. 4 demonstrated INH (Isoniazid) mono-resistance. 12 polydrug resistance demonstrated: 5MDR, 3pre-XDR, while RIF + FQ (fluoroquinolones), FQ + Lz (linezolid), only SLID (second-line injectable drugs), and only FQ resistance observed in 1 case each. Isolated RIF (Rifampicin) resistance and XDR pattern were not observed. Overall frequency of RIF resistance was 16.4% (9/55) and INH was 25% (12/48) with low-(n-2) and high-level INH resistance (n-10). Among second-line drugs, FQ resistance was more than SLID resistance and within FQ, levofloxacin resistance was more frequent than moxifloxacin. MGIT demonstrated positive growth in 16/59 samples, out of which 1 sample was positive for nontuberculous mycobacteria (M. chelonae) but on genotypic testing demonstrated MTB resistant to RIF and FQ. This is the first report on incidence and drug-resistant pattern in culture-positive/negative cases. High (25.49%) primary drug resistance is worrisome. This being the first study in spinal TB cases which document prevalent drug-resistant pattern as evaluated for consecutive culture-positive/negative cases. The tissue obtained must be submitted for AFB culture and molecular tests to ascertain drug resistance in culture-positive/negative cases. However, in the presence of insufficient tissue sample histology and CBNAAT can ascertain etiological diagnosis in 100% cases. INH resistance is more than RIF with isolated RIF resistance unreported.

Genetic surveillance and outcomes of pyrazinamide and fluoroquinolones-resistant tuberculosis in Taiwan

BackgroundPyrazinamide (PZA) and fluoroquinolone (FQ), particularly moxifloxacin (MXF), are essential drugs in the World Health Organization (WHO) recommended short-course regimen to treat drug-susceptible tuberculosis (TB). MethodsTo understand the extent of PZA and MXF susceptibility in general TB cases in Taiwan, we conducted retrospective analyses of 385 conservative Mycobacterium tuberculosis complex (MTBC) isolates identified from 4 TB laboratories in different regions of Taiwan. The case information was obtained from the TB registry. Genotypic drug susceptibility testing (DST) was performed by sequencing drug-resistance associated genes, PZA (pncA) and FQ (gyrA, and gyrB). Phenotypic DST was determined using the Bactec MGIT 960 system or the agar proportion method. Genotyping was carried out using spacer oligonucleotide typing. ResultsIn this study, 4.7% (18/385) cases' isolates harbored pncA mutations and 7.0% (27/385) cases’ isolates harbored gyrA or gyrB mutation. Notably, pncA mutation was associated with Beijing family genotypes (P = 0.028), East African-Indian (EAI) genotypes (P = 0.047) and MDR-TB (P < 0.001). Whereas, gyrA or gyrB mutation was associated with EAI genotypes (P = 0.020) and MDR-TB (P = 0.006). In addition, a statistically significant difference was found between the favorable outcomes using active and inactive PZA (P = 0.009) in 38 case isolates with any pncA, gyrA, or gyrB mutation. ConclusionWe concluded that routine PZA and FQ susceptibility tests are recommended for guiding the treatment of TB.

HPV prevalence and genotype distribution among 38 056 women in Weifang, China: a cross-sectional study

ObjectivesHuman papillomavirus (HPV) infection is closely associated with cervical cancer, especially the persistent infection of high-risk HPV (HR-HPV) genotypes. Therefore, investigating the HPV prevalence, age-specific, genotype distribution and the impact...

Bringing precision medicine to the management of pregnancy in women with glucokinase-MODY: a study of diagnostic accuracy and feasibility of non-invasive prenatal testing.

In pregnancies where the mother has glucokinase-MODY (GCK-MODY), fetal growth is determined by fetal genotype. When the fetus inherits a maternal pathogenic GCK variant, normal fetal growth is anticipated, and insulin treatment of maternal hyperglycaemia is not recommended. At present, fetal genotype is estimated from measurement of fetal abdominal circumference on ultrasound. Non-invasive prenatal testing of fetal GCK genotype (NIPT-GCK) using cell-free DNA in maternal blood has recently been developed. We aimed to compare the diagnostic accuracy of NIPT-GCK with that of ultrasound, and determine the feasibility of using NIPT-GCK to guide pregnancy management. We studied an international cohort of pregnant women with hyperglycaemia due to GCK-MODY. We compared the diagnostic accuracy of NIPT-GCK with that of measurement of fetal abdominal circumference at 28 weeks' gestation (n=38) using a directly genotyped offspring sample as the reference standard. In a feasibility study, we assessed the time to result given to clinicians in 43 consecutive pregnancies affected by GCK-MODY between July 2019 and September 2021. In terms of diagnostic accuracy, NIPT-GCK was more sensitive and specific than ultrasound in predicting fetal genotype (sensitivity 100% and specificity 96% for NIPT-GCK vs sensitivity 53% and specificity 61% for fetal abdominal circumference 75th percentile). In terms of feasibility, a valid NIPT-GCK fetal genotype (≥95% probability) was reported in all 38 pregnancies with an amenable variant and repeated samples when needed. The median time to report was 5 weeks (IQR 3-8 weeks). For the 25 samples received before 20 weeks' gestation, results were reported at a median gestational age of 20 weeks (IQR 18-24), with 23/25 (92%) reported before 28 weeks. Non-invasive prenatal testing of fetal genotype in GCK-MODY pregnancies is highly accurate and is capable of providing a result before the last trimester for most patients. This means that non-invasive prenatal testing of fetal genotype is the optimal approach to management of GCK-MODY pregnancies.

The study of the phenotyping process of some Romanian cherry genotypes

The Plum pox virus (PPV), is considered to be particularly serious in stone species, has appeared in research and documents since 1915 from which revelations emerge about the first symptoms appearing in orchards in central and eastern Europe, where the Plum pox virus (PPV) being so viral, it caused serious production losses in species such as plum, nectarine, cherry, almond, sour cherry, nectarine and last but not least apricot, where losses were 100%. The creation of new varieties with natural genetic resistance to Plum pox virus remains one of the most solid ways to limit the spread of this quarantine virus. The Elisa serological technique and the molecular PCR technique offer the possibility of highlighting the presence or absence of the virus in the plant. The present study illustrates the results of phenotyping tests (indexing on different rootstocks, Elisa and PCR) for some autochthonous cherry genotypes. Even if it is difficult to establish a definitive hypothesis regarding the genetic control of resistance to Plum pox virus in cherry, we still observe an important number of graft/rootstock combinations, such as: Margarit / Otesani 11, Mari de Malu / Otesani 11, Rosu de Mai/ Otesani 11, which showed a resistance to the artificial infection with Plum pox virus. These genotypes are of interest in future breeding works.

Investigating the breeding habitats and kdr allele frequency of malaria vectors in Anambra Central Senatorial Zone, Southeast Nigeria

Malaria, transmitted by infected female Anopheles mosquitoes, is a parasitic disease of major public health importance in Nigeria. Various classes of insecticides have played significant role in the control and elimination of malaria by attacking the vectors. Unfortunately, the success of the use of insecticides in the control of malaria vectors is threatened by the emergence of insecticidal resistance. This study was therefore aimed at determining the resistance status of Anopheles gambiae s. l. in communities in Anambra State, southeast Nigeria. Larval samples were collected from diverse habitats and reared to adult for susceptibility test. Target site resistance assays were carried out and genotypic differentiation test was performed to assess variability of the allelic frequencies of the kdr (L1014F) mutation. The results showed that Anopheles gambiae was the predominant vector (54.2%), followed by Anopheles coluzzii (45.8%). The distribution of the frequency of resistant allele of the kdr gene F (L1014F) showed that the number of homozygous resistant genotype (RR) in Awka North LGA for An. gambiae and An. coluzzii species was highest (31), followed by Awka South (20), and Njikoka (5) LGAs. Also, the number of Heterozygous resistant genotype (Rr) was highest in Awka South (7), followed by Awka North (2), and none was recorded in Njikoka LGA. Homozygous resistant genotype (RR) was frequently observed in the three study areas of Anambra State and is a threat to the use of pyrethroids for insecticidal control of malaria vectors. There is an urgent need for implementation of insecticide resistance management and monitoring in Anambra State to assess the spread of resistance which may expand malaria vector resistance to the remaining classes of insecticides (organophosphates and carbamates) on the long run.