Swab Technique Research Articles

Standardization and validation of ATP luminometry as a diagnostic tool to assess the cleanliness of feeding equipment in preweaning calves

The objective of this cross-sectional study was to standardize a reliable and repeatable swabbing technique using ATP luminometry (light emission proportional to the amount of ATP with result provided in relative light units [RLU]) to describe the cleanliness of various feeding equipment used for preweaning calves in dairy farms. A total of 7 Québec commercial dairy herds were selected conveniently. Following visual hygiene scoring, the cleanliness of every available piece of feeding equipment was assessed using direct surface swabbing for buckets and nipples with Hygiena UltraSnap swabs. A liquid rinsing technique was used for esophageal feeders, bottles, and automatic milk feeders (AMF) with UltraSnap, AquaSnap, and MicroSnap swabs. To validate direct swabbing technique of buckets, a stage within and between operators was realized, as well as a conventional bacterial culture. A total of 519 swab samples were obtained from 201 pieces of equipment. The median (interquartile range) contamination in RLU for a bottle, esophageal feeder, AMF, bucket and nipple was 2 (1;6), 2 (0;12), 52 (19;269), 886 (128;7,230) and 899 (142;6,928), respectively. The direct swabbing technique, which consists in swabbing directly the surface of an equipment, showed excellent correlation for intrarater reliability (intraclass correlation (ICC) = 0.93; 95% CI: 0.88-0.96). The interoperator (2 sessions with 3 different operators) reliability also showed high correlation (ICC = 0.88; 95% CI: 0.78-0.94 for the first session, and ICC = 0.89; 95% CI: 0.79-0.95 for the second session). Luminometer values were positively associated with the visual score of esophageal feeders, AMF and buckets. A positive correlation between bacterial culture and direct swabbing of buckets was also found for the UltraSnap (rs = 0.653; 95% CI: 0.283-0.873; P = 0.0003) and MicroSnap (rs = 0.569, 95% CI: 0.309-0.765; P = 0.002). This study describes a standardized and practical on-farm swabbing technique for assessing the hygienic status of feeding equipment by luminometry, which can be integrated in the investigation of preweaning dairy calves problems.

Pathogenic Bacterial Communities Isolated and Identified in Stingless Bee (Kelulut) Honey from Selected Farms in Terengganu

In Malaysia, stingless bees can be categorised into two genera: Melipona and Trigona, known as “kelulut”. The high demand for kelulut honey boosts the production of the honey industry. Previous studies reported that stingless bee (kelulut) honey and its products were contaminated with pathogenic bacteria during harvesting and processing. This research aims to isolate and identify the pathogenic bacteria in kelulut honey. Forty-eight samples of kelulut honey (open and closed pot) and propolis were obtained from selected farms in Terengganu by focusing on a major stingless bee species available in Malaysia, Heterotrigona itama. In addition, the swabbing technique was done on the wooden beehive of the kelulut to evaluate the environmental contamination. The pathogenic bacteria were isolated using specifically selected agar, such as Bacillus cereus agar (for B. cereus), Baird-Parker agar (for Staphylococcus aureus), and MacConkey agar (for other pathogenic bacteria), which were confirmed through a biochemical test. All samples were analysed, and the results showed that B. cereus (7/48), Pseudomonas aeruginosa (10/48), Pantoea spp. (11/48), Serratia plymuthica (6/48), and S. aureus (9/48) were obtained in the samples. This study indicates that kelulut honey was contaminated with B. cereus, P. aeruginosa, Pantoea spp., S. plymuthica, and S. aureus. Isolated pathogenic bacteria may exist in the kelulut honey through food handlers, utensils, and the environment. Hence, the stakeholders should strictly follow good standard operating procedures and guidelines by the kelulut honey industry to prevent foodborne illness.

Human skin biofilm model: translational impact on swabbing and debridement

Objective: Wound biofilms are one of the greatest challenges in the therapy of hard-to-heal (chronic) wounds, as potent antimicrobial substances fail to eradicate bacteria within short incubation periods. Preclinical investigations using novel model systems that closely mimic the human wound environment and wound biofilm are required to identify new and effective therapeutic options. This study aims to identify bacterial colonisation patterns that are relevant for diagnosis and therapy. Method: In this study, a recently established human plasma biofilm model (hpBIOM) was incorporated into a wound within human dermal resectates after abdominoplasty. The interaction of the biofilm-forming bacteria meticillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa with the skin cells was investigated. Possible effects on wound healing processes in correlation with the persistence of the biofilm in the wound environment were analysed in patients with leg ulcers of different aetiologies and biofilm burden. Results: Using haematoxylin and eosin staining, species-dependent infiltration modes of the bacteria into the wound tissue were determined for the pathogens MRSA and Pseudomonas aeruginosa. The spreading behaviour correlated with clinical observations of the spatial distributions of the bacteria. In particular, the clinically prominent Pseudomonas aeruginosa-specific distension of the wound margin was identified as epidermolysis due to persistent infiltration. Conclusion: The hpBIOM applied in this study represents a potential tool for preclinical analyses dealing with approval processes for new antimicrobial applications. In terms of clinical practice, a microbiological swabbing technique including the wound margin should be routinely applied to prevent wound exacerbation.

Success of different therapy concepts for bacterial endometritis in stud farm practice

Bacterial endometritis is among the major problems in equine reproduction and is usually treated with antibiotics. Data on clinical success rates of antibiotic as well as non-antibiotic treatment is scarce. The study aimed to collect data of mares diagnosed with intrauterine growth of facultative pathogenic bacteria and compare the outcome of different therapies for bacterial endometritis in stud farm practice. Data on mares (different breeds) with positive uterine culture results was collected retrospectively in private veterinary practices (n=5) from 2018 to 2022. Information relating to 30 factors (mare, diagnostics, therapy and pregnancy rate) of cases with bacterial endometritis (Endometritis, n=1005) were recorded and analysed. Possible effects on treatment success (negative control swab or positive pregnancy result in the first heat after treatment) were tested by binomial logistic regression. For comparison of pregnancy rates, data on mares with negative uterine culture and without any treatment (Control; n=446) was also collected and compared with Endometritis group by chi square testing. In group Endometritis, uterine swabbing (technique: 72% double-guarded swab, 28% low-volume lavage) revealed presence of β-hemolytic streptococci in most cases (n=918). Pseudomonas aeruginosa (n=21), Klebsiella pneumoniae (n=15), Staphylococcus aureus (n=13) and Candida sp. (n=10) were observed less frequently. Results positive for E. coli were not included. In group Endometritis, treatment occurred regardless of the cycle stage either with trimethoprim-sulfonamides (n=532), procain-penicillium (n=277),marbofloxacin (n=71), other antibiotics (n=66) or without antibiotics (n=59) after susceptibility testing (performed in 549/1005 cases). Antibiotics were administered rather intrauterine (n=34) than systemically (n=911). Uterine lavage was reported in 46% of mares. Additional local treatment with different substances (e.g., n-acetylcysteine, dimethyl sulfoxide or Coca Cola) was performed in 164 mares. Uterotonic drugs were administered in 39.9% of mares. A control uterine culture was not obtained in most cases (n=677). Different breeding programs in this study included artificial insemination (AI) with chilled semen (n=667), AI with frozen semen (n=169), transfer of fresh (n=112) or cryopreserved (n=27) embryos and natural cover (n=27). In the control group, AI with chilled semen (n=230), AI with frozen semen (n=106), embryo transfer (fresh: n=49; cryopreserved: n=58) and natural cover (n=2) were used. After the first breeding after treatment, pregnancy rate was 47% in group Endometritis and 54% in the control group (p<0.05). End of season pregnancy rate was 72% in treated mares and 76% in the control group (p>0.05). Treatment success was affected significantly by duration of antibiotic treatment, usage of uterotonic drugs, veterinary practice, and mare's age, but neither by the chosen antibiotic drug nor administration route nor covering method. In conclusion, end of season pregnancy rate did not differ among mares treated for bacterial endometritis vs. mares with negative uterine cultures.

Biophysical indicators of the mucous membrane of the oral cavity, the microcirculatory channel and the oral fluid in patients with lichen ruber planus

Annotation. The work aims to conduct a biophysical assessment of the charge state of the buccal epithelium cells (BEC), the microcirculatory channel of the oral mucosa and oral fluid in patients with the erosive-ulcerative form of lichen planus. The problem is relevant due to the increasing number of lichen planus patients, the uncertain etiology of the disease, and the ongoing struggle to find treatment solutions. 11 patients aged 50-60 years with lichen planus (erosive-ulcerative form) were examined. All patients underwent a study of the charge state of the buccal epithelium cells. Buccal cell samples were collected on an empty stomach after rinsing the oral cavity with water, applying a gentle swabbing technique. A spectrocolorimeter of type “Pulsar” was used to assess the condition of the microcapillary channel of the oral mucosa. In addition, in patients with lichen planus, the stability of the pH of the oral fluid (ΔpH) was evaluated as one of the indicators of the level of non-specific resistance in the body and the oral cavity in particular. Five 1ml samples of oral fluid were taken from each patient to calculate the value of ΔpH, where pH value was determined using an ionometer immediately after sampling. An average value of ΔрН and the confidence interval of deviations (ΔрН) from the average value were calculated, taking into account the Student coefficient for five measurements and a confidence probability of 0.95. The results obtained from biophysical studies show a significant decrease in all studied indicators. The percentage of mobile nuclei in patients with the erosive-ulcerative form of lichen planus of the oral mucosa is two times lower than in healthy people of the same age. At the same time, the plasmalemma and nuclei displacement amplitudes, as well as their ratio, were reduced compared to the norm, which indicates a decrease in their charge and deterioration of the physiological state of cells and the body as a whole. We found that individuals with lichen planus exhibit a decreased efficiency of functional reactions in the oral cavity tissues, which requires the development and implementation of effective treatment and prevention measures.

Relative Abundance and Detection of Pseudomonas aeruginosa from Chronic Wound Infections Globally.

Pseudomonas aeruginosa is a difficult-to-treat pathogen that is frequently involved with chronic wound infections. Here, we conducted a literature search of world-wide studies published between 2005 and 2022 that described the microbiological profiles of chronic wound infections. For each continent, a hierarchy of pathogens was created to define the organisms that were most frequently isolated in each region. Except for South America, P. aeruginosa was the second most common organism in each major continent, with Staphylococcus aureus being the most abundant pathogen overall. When individual countries were evaluated, P. aeruginosa was the most frequently isolated organism in several Southeast Asia nations including India and Malaysia. P. aeruginosa was less commonly isolated from diabetic foot infections in North America, Europe, and Africa in comparison to other types of chronic wound infections. Additionally, the Levine wound swab technique may be a quick and painless way to isolate P. aeruginosa from wound infections, but the isolation of P. aeruginosa does not seem to be an informative predictor of the patient's clinical course. A multivariate risk assessment that accounts for the regional frequency of P. aeruginosa isolation may be an appropriate way to guide empiric management of chronic wound infections.

Selective detection of urease-producing bacteria on the genital skin surface in patients with incontinence-associated dermatitis.

We aimed to investigate the association between the presence of cutaneous urease-producing bacteria and the development of incontinence-associated dermatitis (IAD) using an original urea agar medium as a step toward developing advanced preventive measures. In previous clinical assessments, we developed an original urea agar medium to detect urease-producing bacteria via the medium's colour changes. In a cross-sectional study, specimens were collected via the swabbing technique at genital skin sites in 52 stroke patients hospitalised in a university hospital. The primary objective was to compare the presence of urease-producing bacteria between the IAD and no-IAD groups. Determining the bacterial count was the secondary objective. The prevalence of IAD was 48%. A significantly higher detection rate of urease-producing bacteria was observed in the IAD group than in the no-IAD group (P = .002) despite the total number of bacteria being equivalent between them. In conclusion, we discovered that there was a significant association between the presence of urease-producing bacteria and IAD development in hospitalised stroke patients.

Intermediate Cluster Disinfection: Which Disinfection Solution Is Most Effective on Milking Liners? A Comparison of Microorganism Reduction on Liner Inner Surfaces Using Quantitative Swab Sampling Technique.

During machine milking, pathogenic microorganisms can be transmitted from cow to cow through liners. Therefore, in Germany, a spray method for the intermediate disinfection of the milking cluster is often used for prevention. This method of cluster disinfection is easy to perform, requires little time and no extra materials, and the disinfection solution is safe from outside contamination in the spray bottle. Since no data on a systematic efficacy trial are available, the aim of this study was to determine the microbial reduction effect of intermediate disinfection. Therefore, laboratory and field trials were conducted. In both trials, two sprays of 0.85 mL per burst of different disinfectant solutions were sprayed into the contaminated liners. For sampling, a quantitative swabbing method using a modified wet-dry swab (WDS) technique based on DIN 10113-1: 1997-07 was applied. Thus, the effectiveness of disinfectants based on Peracetic Acid, Hydrogen Peroxide and Plasma-Activated Buffered Solution (PABS) was compared. In the laboratory trial, the inner surfaces of liners were contaminated with pure cultures of Escherichia (E.) coli, Staphylococcus (S.) aureus, Streptococcus (Sc.) uberis and Sc. agalactiae. The disinfection of the contaminated liners with the disinfectants resulted in a significant reduction in bacteria with values averaging 1 log for E. coli, 0.7 log for S. aureus, 0.7 log for Sc. uberis and 0.8 log for Sc. agalactiae. The highest reduction was obtained for contamination with E. coli (1.3 log) and Sc. uberis (0.8 log) when PABS was applied and for contamination with S. aureus (1.1 log) and Sc. agalactiae (1 log) when Peracetic Acid Solution (PAS) was used. Treatment with sterile water only led to an average reduction of 0.4 log. In the field trial, after the milking of 575 cows, the liners were disinfected and the total microorganism count from the liner surface was performed. The reduction was measured against an untreated liner within the cluster. Although a reduction in microorganisms was achieved in the field trial, it was not significant. When using PAS, a log reduction of 0.3 was achieved; when using PABS, a log reduction of 0.2 was obtained. The difference between the two disinfection methods was also not significant. Treatment with sterile water only led to a reduction of 0.1 log. The results show that spray disinfection under these circumstances does result in a reduction in the bacteria on the milking liner surface, but for effective disinfection a higher reduction would be preferred.

Abstract 2375: Whole exome sequencing and immune profiling of cervical adenocarcinoma reveal a “cold” tumor-immune phenotype

Abstract Purpose: Cervical adenocarcinomas (CAC), a less common cervical cancer histology with poor prognosis as compared to squamous cell carcinoma (CSCC), disproportionally present at advanced stages in young women and incidence is increasing; however, the same chemoradiation (CRT) paradigm is used by clinicians for both histologies. Given the rarity of CAC, little is known about genomic pathways and tumoral immune factors that differentiate CAC from CSCC and could drive differences in patient outcomes. Objective: To identify distinctions in the mutational landscape and immune phenotype of CAC and CSCC tumors in patients undergoing CRT which may be potential therapeutic targets. Methods: 80 patients with FIGO stage IB-IVA cervical cancer who underwent definitive CRT were prospectively enrolled on an IRB approved clinical trial. Cervical tumor DNA (9 CAC patients; 58 CSCC patients) and cervical tumor microenvironment samples (13 CAC patients, 67 CSCC patients) were collected by a noninvasive swab technique at baseline, and after 1, 3, and 5 weeks of CRT. WES and neoantigen prediction were performed and tumor mutation burden (TMB) was calculated as the number of somatic coding mutations per megabase. Lymphocyte immunostaining was performed according to standard protocols and cells were analyzed using a LSRFortessa X-20 Flow Cytometer (BD Biosciences) and FlowJo 10.6.1. Statistical comparison of mean values was performed by t-test. Results: Baseline mean TMB and tumor neoantigen was lower in CAC compared to CSCC (1.1±0.14 vs 5.2±3.5 respectively for TMB, p&amp;lt;0.01; 6.0±8.5 vs 455.3±472.5 respectively for neoantigen, p&amp;lt;0.01) and persisted during CRT. The top 3 most frequently altered genes unique to CAC were GPR98, SLC12A3, and TP53. The baseline mutational profile of CAC tumors was most similar to other adenocarcinomas including prostate and pancreatic cancer in TCGA. There was a significant reduction for CAC in baseline CD4+ T-cell infiltration in CAC compared to CSCC (p=0.01) which persisted through the course of CRT (p=0.02 at week 3). There was a trend toward decreased CD4+ T-cell activation, including co-expressing CD69 (p=0.17) and Ki67 (p=0.06), decreased CD8+ T-cell infiltration (p=0.19), and decreased Ki67+CD8+ T-cells as compared to CSCC (p=0.02). Conclusions: In contrast to CSCC, immune profiling of the CAC tumor microenvironment suggests a reduction in immune cell infiltrate and absence of or failed T-cell activation. Similarly, unique genetic alterations were detected between histologies, and CAC tumors displayed reduced TMB and loss of tumor neoantigen expression. These findings are consistent with an “immune cold” phenotype in CAC. These patients may be less likely to benefit from immunotherapy. Future investigation into molecularly targeted therapies or strategies that help mobilize peritumoral T-cells in response to CRT may better potentiate improved CRT response in CAC. Citation Format: Julianna Bronk, Xiaogang Wu, Rui Wang, Ananta Yanamandra, Tatiana Karpinets, Chike Abana, Molly El Alam, Katarina Tomasic, Daniel Lin, Erica Lynn, David Lo, Timothy Harris, Xingzhi Song, Andrew Futreal, Jianhua Zhang, Jagannadha Sastry, Ann Klopp, Lauren Colbert. Whole exome sequencing and immune profiling of cervical adenocarcinoma reveal a “cold” tumor-immune phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2375.

Effect of swabbing technique and duration on forensic DNA recovery

Various factors have been shown to affect performance of the conventional wet-dry double and single wet swabbing techniques to recover DNA, such as pressure and angle of application, volume and type of wetting agent, and swab type. However, casework laboratories in some jurisdictions have recently adopted different swabbing techniques that include wet-moist double swabbing and moist-dry single swabbing. Factors affecting the effectiveness of these recent techniques in maximising DNA recovery therefore need to be investigated. Here, the performance of traditional and recent swabbing techniques was compared and the impact of swabbing duration on DNA recovery was investigated. Ten µl aliquots of a known concentration of DNA extracted from human blood were deposited on pre-cleaned DNA-free cotton swatches (porous) and porcelain tiles (non-porous). Five swabbing techniques were used, of which three were double swabbing techniques: wet-moist, wet-wet and wet-dry, and two were single swabbing techniques: wet and moist-dry. For a ‘wet’ or ‘moist’ swab, 100 or 50 µL water was added, respectively. For a moist-dry swab, water was applied to one side of the swab, leaving the other side drier. Each swabbing technique was applied for two durations, 15 and 30 s per swab, with 5 reps of each combination (n = 100 plus controls). All samples were extracted and quantified, and a sub-set was profiled. The results showed that the wet-moist double swabbing technique with a swabbing duration of 30 s maximised DNA recovery from cotton. From tile, a single wet or moist-dry swab maximised DNA recovery, but increasing swabbing duration from 15 to 30 s had no impact. These data can be used to inform standardisation of DNA collection protocols across casework laboratories.

UJI DAYA HAMBAT EKSTRAK Cymbopogon citratus DAN Alpinia purpurata TERHADAP PERTUMBUHAN Shigella sonnei

This study aimed to determine the phytochemical content and potency of the ethanol extract of lemongrass (Cymbopogon citratus) stem and red galangal rhizome (Alpinia purpurata) in inhibiting the growth of Shigella sonnei in vitro. Phytochemical testing of ethanol extract using Thin Layer Chromatography (TLC). Antibacterial test was carried out by applying the Kirby Baeur disk diffusion method with swab technique with a series treatment of extract concentrations of 6.25%, 12.5%, 25%, 50%, positive control (tetracycline 20%), and negative control (DMSO 10%) with 5 replicates. From the results of phytochemical tests, showed that secondary metabolites of flavonoids and terpenoids were found in red galangal extract. While the lemongrass extract showed positive results for alkaloids, flavonoids, terpenoids and phenolic compounds. The ethanol extract of lemongrass stem and galangal rhizome potential as an antibacterial with the highest inhibitory activity obtained at a concentration of 50% in both lemongrass and galangal extracts, respectively 8.4 mm and 7 mm. The results of the antibacterial test showed that red galangal extract and lemongrass extract had phytochemical contents that could inhibited the growth of S. sonnei.

A Study on Effects of Immersion in River Water and Sea Water on DNA Placed on Guns

To determine the quantity of DNA that can be obtained from that placed on guns after 30 minutes, 90 minutes and 270 minutes of immersion in water. Also, the aim of this study was to disclose how different types of water environments, such as freshwater and saltwater, affect the amount of DNA loss and degradation over the preset period of time.&#x0D; Firearms was swabbed with a double swabbing technique. DNA was isolated using a Qiagen procedure, quantified by real-time PCR, amplification was on thermal cycler and Capillary electrophoresis was performed on 3500 Genetic analyzer. The electropherograms obtained were analyzed with the GeneMapper ID-X software.&#x0D; Through the comparison of DNA amounts detected after isolation and DNA profiles/allele numbers detected on guns, it can be concluded that the influence of saltwater and freshwater are significantly different. The loss of DNA in saltwater from the outer firearms surface after 30 min is similar to the loss of DNA in freshwater after 90 minutes of immersion.&#x0D; Based on the results of this study, especially the results of real casework sample, it can be concluded as expected that saltwater has a more serious effects on the degradation and the loss DNA on guns than freshwater. Further, it point out that the guns found in freshwater after crime could be a valuable piece of forensic evidence, whilst the guns recovered from saltwater will have less value as forensic evidence if found after more than 30 minutes

Feasibility of a novel non-invasive swab technique for serial whole-exome sequencing of cervical tumors during chemoradiation therapy.

BackgroundClinically relevant genetic predictors of radiation response for cervical cancer are understudied due to the morbidity of repeat invasive biopsies required to obtain genetic material. Thus, we aimed to demonstrate the feasibility of a novel noninvasive cervical swab technique to (1) collect tumor DNA with adequate throughput to (2) perform whole-exome sequencing (WES) at serial time points over the course of chemoradiation therapy (CRT).MethodsCervical cancer tumor samples from patients undergoing chemoradiation were collected at baseline, at week 1, week 3, and at the completion of CRT (week 5) using a noninvasive swab-based biopsy technique. Swab samples were analyzed with whole-exome sequencing (WES) with mutation calling using a custom pipeline optimized for shallow whole-exome sequencing with low tumor purity (TP). Tumor mutation changes over the course of treatment were profiled.Results216 samples were collected and successfully sequenced for 70 patients (94% of total number of tumor samples collected). A total of 33 patients had a complete set of samples at all four time points. The mean mapping rate was 98% for all samples, and the mean target coverage was 180. Estimated TP was greater than 5% for all samples. Overall mutation frequency decreased during CRT but mapping rate and mean target coverage remained at >98% and >180 reads at week 5.ConclusionThis study demonstrates the feasibility and application of a noninvasive swab-based technique for WES analysis which may be applied to investigate dynamic tumor mutational changes during treatment to identify novel genes which confer radiation resistance.

Comparison of operational DNA recovery methods: Swabs versus tapelifts

It is routine among many jurisdictions to recover DNA using tapelifts on porous substrates (e.g. clothing) and swabs on non-porous substrates (e.g. tool handles). Here, we examine this by comparing the efficiency of the NSW jurisdiction’s specific swabbing and tapelift techniques on a range of porous and non-porous substrates. To test DNA recovery efficiency, 30 μl aliquots of 1:50 and 1:100 saliva dilutions were deposited onto the substrates, left to dry overnight, recovered, extracted, quantified and a subset profiled. Tapelifts recovered more DNA and DNA profiles with more detectable alleles than swabs for both saliva dilutions on porous substrates. For non-porous substrates, similar DNA quantities and profiles were generally recovered with both methods for both saliva dilutions. These data underpin current practices to recover DNA using tapelifts for porous substrates and swabs for non-porous substrates. These data also revealed severe degradation of DNA recovered from brass, supporting the on-going need to improve DNA recovery and analysis methods for brass substrates.

Isolation and characterization of uterine leukocytes collected using a uterine swab technique.

Leukocytes from the maternal-fetal interface are a valuable tool to study local changes in immune function during pregnancy; however, sampling can be challenging due to inadequate tissue availability and the invasive nature of placental bed biopsy. Here, we aim to purify and characterize leukocytes from paired peripheral and uterine blood samples to assess whether a less invasive method of uterine blood collection could yield a population of enriched uterine leukocytes suitable for ex vivo and in vitro analyses. Human peripheral blood mononuclear cells (PBMC) and uterine blood mononuclear cells (UBMC) expressed from surgical gauze post C-section were isolated, and immunophenotypic information was acquired by multi-parameter flow cytometry. PBMC and UBMC were stained for markers used to define T and B lymphocytes, macrophages, regulatory T (TReg ) cells, and natural killer (NK) cells. Prime flow was performed to check expression and analysis of CD16- CD56++ and CD16- CD56++ NK transcripts in PBMC and UBMC samples. Immunophenotyping revealed that over 95% of both live PBMC and UBMC consisted of CD45+ leukocytes. Higher percentages of CD16- CD56++ , characterized as uterine NK (uNK) cells, were observed in UBMC samples as compared to PBMC samples (18.41% of CD45+ CD3- vs. 2.73%, respectively), suggesting that CD16- CD56++ cells were enriched in these samples. In UBMC, 49.64% of CD3-negative cells were of peripheral NK phenotype (CD16+ CD56++ ), suggesting infiltration of maternal peripheral NK (pNK) cell in the uterine interface. Intrauterine leukocytes, especially CD16- CD56++ NK cells, can be collected in sufficient numbers with increased purity by sampling the uterine cavity postdelivery with surgical gauze. Our results suggest that this non-invasive protocol is a useful sampling technique for isolating CD16- CD56++ cells, however, due to peripheral blood contamination, the NK cell yield could be lower compared to actual decidual or endometrial samples post-partum which is more invasive.

Infertility In Queens: Clinical approach, experiences and challenges.

It is not easy for a veterinarian to determine the cause of infertility in a queen. The simplest method is to start by investigating the most common causes and then progress by successive elimination to the less common causes. For example, mating problems are commonly responsible for reproductive failure, and owners should be particularly vigilant in checking that the mating is progressing satisfactorily. Frequently, a veterinarian may have to determine, among other things, if there is a uterine or hormonal disorder or if an infectious pathology may be involved. In the case of a queen that is not cycling, it is important to differentiate a lack of onset of puberty in a young queen from a lack of cyclicity in an adult queen. This review sets out a practical, step-by-step approach that veterinarians can follow, starting with the most common causes of infertility in queens and moving towards more rare or specific causes. Simple techniques such as vaginal smears are essential to determine if a queen that is ready to be mated is in full oestrus, if a queen is cycling or not and to diagnose inflammation of the genital tract. Veterinarians should be familiar with the peculiarities of the swabbing technique in cats, as well as the interpretation of feline vaginal cytology. Progesterone assays are useful on many occasions in the diagnosis pathway. A laboratory capable of performing karyotypes should ideally be available. Finally, it is important to be able to perform an ultrasound examination of the ovaries and uterus in a queen. Although it appears that infertility is a problem among purebred queens, there is a lack of knowledge in this area and too few published studies. Some causes of feline infertility remain poorly investigated.

Lab practices that improve coronavirus disease 2019 detection accuracy using real-time PCR.

ABSTRACTThe number of coronavirus disease 2019 (COVID-19) cases significantly increased with the emergence of multiple variants of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). This has led to an ongoing effort focused on developing the diagnostic detection tests. Among the currently available tests, real-time reverse transcriptase PCR (RT-PCR) has been considered as the ‘golden method’ for the detection of SARS-COV-2. However, a significant number of inaccurate (false-negative/false-positive) results have been reported in spite of this method's reliability and effectiveness. These unreliable results may arise because of various issues encountered throughout the entire testing process starting with the sampling phase, going through the PCR process, and ending with the result analysis. This article aims to shed light on the errors that occur during the COVID-19 testing process and suggest ways to overcome them effectively. Accurate testing could be optimized by following the correct swabbing technique, using adequate RT-PCR kits and controls, setting clear lab guidelines, and properly interpreting the results.

Molecular Identification of Biofilm-forming Pseudomonas aeruginosa Isolates from Noncritical Surfaces of a Tertiary Healthcare Center in Abia State, Southeast Nigeria

The presence of biofilm-forming Pseudomonas aeruginosa on noncritical surfaces in a hospital has been attributed to healthcare-associated infections (HAIs) leading to multidrug-resistant (MDR) infections. This study is aimed at identifying the molecular characteristics of Biofilm forming Pseudomonas aeruginosa found on non-critical surfaces in a hospital environment. Samples were collected using the swab technique from different noncritical surfaces surrounding hospitalized patients in different wards namely sphygmomanometers, thermometers, bed rails etc. Bacterial analysis was performed by using conventional microbiological techniques, biochemical tests and Microbact 24E assay. Biofilm forming Pseudomonas aeruginosa were visualized using Crystal Violet staining assay and then estimated by measuring the Optical Density through Spectrophotometer. Four hundred and fifty (450) positive samples of bacterial isolates from noncritical surfaces of six selected wards of the hospital were obtained from one thousand three hundred and fourteen (1314) samples. One hundred and thirty-nine (139) isolates of Pseudomonas spp. were obtained, out of which 115 (82.73%) were identified as biofilm formers. Of these 115 biofilm formers, 83(72.17%) belonged to Pseudomonas aeruginosa, 30(22.09) Pseudomonas fluorescens, 20(17.39%) Pseudomonas putida and 6(5.22%) belongs to Pseudomonas chlorophis. The final characterization of Biofilm forming Pseudomonas aeruginosa isolates was verified by 16S rDNA sequencing. Sequence analysis showed similarity at 99.5% Pearson`s correlation coefficient (r) between 16S rDNA of the representative standard isolate (from gene library) and Biofilm forming Pseudomonas aeruginosa strain obtained from the Tertiary health facility, Federal Medical Centre (FMC), Umuahia. The nucleotide sequence of the experimental isolates expressed genetic mutation which represents the point of involvement of Pel B gene and is involved in the biofilm formation character of Pseudomonas aeruginosa in this study. There is an urgent need to take nosocomial transmission from noncritical surfaces serious in FMC Umuahia.

Skin Microbiota Profiles from Tape Stripping and Skin Biopsy Samples of Patients with Psoriasis Treated with Narrowband Ultraviolet B.

PurposeAlthough the pathogenesis of psoriasis involves the dermis, most previous studies collected samples using the swab technique. A recent study examining the microbiomes obtained via both skin biopsies and swabs revealed a significant difference in normal skin. We hypothesized that the microbiome profile of patients with psoriasis from tape stripping and skin biopsy might be different. This study sought to contribute to microbiome research on psoriasis by investigating the changes in the microbiome during narrowband ultraviolet B (NBUVB) therapy by comparing the results from the different sampling techniques of tape stripping and skin biopsy.Patients and MethodsTwenty-three participants, including 14 patients with chronic plaque psoriasis and nine healthy controls, were recruited, and nine patients with psoriasis completed 20-sessions of NBUVB treatment. Skin microbiota from both techniques was analyzed using the 16S rRNA gene at baseline and after treatment.ResultsA clear difference was observed between the results from the two sampling techniques. Alpha diversity of the microbiota obtained from tape stripping was higher than that of the microbiota from skin biopsy, whereas beta diversity was clustered into two groups by sampling technique. The microbiome was altered during NBUVB treatment using both sampling techniques.ConclusionDifferent sampling techniques resulted in different microbiome profiles in patients with psoriasis. Tape stripping and swabs are feasible procedures and are mostly used in psoriasis and other skin microbiome studies; however, skin biopsy may also expand our understanding of psoriasis and other skin diseases that pathophysiology involves deeper to the dermis or subcutaneous tissue.

The Impression of Blood Droplets on Solid Surface and its Evaluation with Bloodstain Pattern Analysis, A Cross Section Forensic Study

Task: The aims and objectives of current study were to identify the values of bloodstain pattern analysis for blood droplets on solid surface in forensic sciences. Methodology: Study design: Present study was conducted in different medical institutions of Lahore from November 2021to March 2022. Crime scene (Blood droplets on solid surface): In current study for research purpose few blood droplets of 2 individuals were dropped on different solid surfaces and consider it as crime scene. Sample collection: In this study total 10 blood samples were collected from different individuals for cross match with the droplets of blood sample collected from solid surfaces. 2, individuals how dropped their blood droplets on solid surfaces were also included in total participants of present study. Blood droplets from solid surfaces were collected with the help of cotton swab techniques. DNA analysis: With the application of Polymerase chain reaction such analysis performed and DNA released from the cell by extraction and measured the amount of genetic material than amplified its copies and separated these amplified copies from each other. The qualitatively and quantitatively comparing of DNA evidence samples are referred as DNA profiles and checking the technical correctness of analyst reports was its quality assurance.