This review focuses on describing new commercially available POC-type molecular diagnostic systems that can be easily implemented in community pharmacies and have the potential to expand the portfolio of pharmaceutical services and make a significant contribution to the improvement of public health.Knowledge of new molecular diagnostic techniques other than PCR is relatively unexplored. However, the available options are diverse and have reached sufficient technological maturity for large-scale use. The SARS-CoV-2 pandemic has brought diagnostic tests to market that, in some cases, have been used exclusively in research for decades.Isothermal nucleic acid amplification technology continues to evolve and it is likely that in the coming years we will witness an exponential increase in its use, as well as the development of new improvements that further simplify and reduce the cost of each assay.Furthermore, we cannot ignore the fact that during the COVID-19 pandemic, the public has become accustomed to self-diagnosing through mass distribution channels such as community pharmacies. Which can open the sector to other diseases - such as sexually transmitted diseases or animal health -, food control, water and air contamination (fungi) or the presence of allergens.Knowledge of them is an essential technological surveillance strategy for the pharmaceutical sector.

Trichomoniasis is a parasitic disease that affects the human reproductive and urinary systems, representing a substantial non-viral sexually transmitted infection worldwide. Given its impact on reproductive health, and the limited available information on the prevalence of Trichomonas vaginalis, this study aimed to evaluate the prevalence of T. vaginalis among women referred to health centers in Tabriz, Northwest Iran. Study was conducted on 448 suspicious women who attended to 29Bahman hospital in Tabriz, Northwest Iran, during September 2020 to September 2021. Demographic data were collected according to the study protocol. Vaginal discharges were obtained using sterile swabs, and the prevalence of T. vaginalis was determined using Papanicolauo staining and PCR method. Among the 448 cases studied, 48 (10.7%) samples were suspected as a T. vaginalis infection, while 4 (0.89%) confirmed using the PCR method. The mean age of infected individuals was 41.7 ± 9.4 years. No statistical correlation was observed between inflammation, method of contraception and infection (p = 0.8). The present study revealed a relatively low prevalence of T. vaginalis infection within the study population. Additionally, the utilization of the PCR method can be beneficial in confirming suspected samples.

Bovine theileriosis is a protozoan disease caused by the intracellular parasite (Theileria spp.) transmitted by ticks and it is considered one of the most significant parasitic diseases, potentially endangering Egyptian cattle herd industry. The present study was conducted for a molecular survey of bovine theileriosis and its associated risk factors (season variations, geographical locations, breeds, age, sex, tick infestation, and acaricide applications) in three Egyptian governorates, Beni-Suef, Al-Faiyum, and Al-Minya for a year extended from December 2021 to November 2022, in addition, genetic diversity of Theileria isolates. A total of 961 cattle were examined for Theileria infection clinically, microscopically, and by Polymerase Chain Reaction (PCR) using 18S rRNA gene for piroplasms DNA detection, Theileria genus-specific primers of the small subunit of rRNA gene, and Theileria annulata specific primers of the Tams-1 gene. The prevalence rate of bovine theileriosis was 9.26%, and 11.86% using Giemsa-stained blood smear and PCR, respectively. All positive samples screened by Theileria genus-specific primers were positive for T. annulata when screened by the specific primers. Based on molecular screening, season, cattle breeds and acaricide applications were considered risk factors for T. annulata infection, while locality, age, sex and tick infestation had insignificant effects with the occurrence of the disease. A potential novel T. annulata haplotype based on the Tam-1 gene was identified with accession numbers OR364144 and OR915851. Therefore, T. annulata was the only Theileria species found and played a significant problem in the cattle population. This study could be the basis for future studies on unexplored regions and different animal species for well-structured prevention and control measures.

Detecting life has driven research and exploration for centuries, but recent attempts to compile and generate a framework that summarizes life features, aimed to develop strategies for life detection missions beyond planet Earth, have disregarded a key life feature: behavior. Yet, some behaviors such as biomineralization or motility have occasionally been proposed as biosignatures to detect life. Here, we capitalize on a specific taxis\' motility behavior, magnetotaxis, to experimentally provide insights in support of behavior as an unambiguous, sensitive biosignature, and magnetic forces as a prescreening option. Using a magnetotactic bacterial species, Magnetospirillum magneticum, we conducted a lab sensitivity experiment comparing PCR with the hanging drop behavioral assay, using a dilution series. The hanging drop behavioral assay visually shows the motility of MTB toward magnetic poles. Our findings reveal that the behavioral assay exhibits higher sensitivity in the detection of M. magneticum when compared to the established PCR protocol. While both methods present similar detection sensitivities at high concentrations, at ≥ 10-7 fold dilutions, the behavioral method proved more sensitive. The behavioral method can detect bacteria even when samples are diluted at 10-9. Comparable results were obtained with environmental samples from the Hula Valley. We propose behavioral cues as valuable biosignatures in the ongoing efforts of life detection in unexplored aquatic habitats on Earth and to stimulate and support discussions about how to detect extant life beyond Earth. Generic and robust behavioral assays can represent a methodological revolution.

[This corrects the article DOI: 10.3389/fvets.2023.1266499.].

Hemophilia B (HB) is an inherited bleeding disorder caused by defects in the FⅨ gene, leading to severe coagulation dysfunction. This study designed eight pairs of primers covering eight exons of the FⅨ gene and used PCR and DNA sequencing to detect FⅨ gene mutations in 31 HB patients. Sequencing results were compared with normal sequences using Chromas software on Blast to identify mutation sites. Findings revealed the CpG dinucleotide region as a mutation hotspot and the 192nd nucleotide (FⅨ192) as a dinucleotide polymorphism site in the Chinese population. Pathogenic mutations included point mutations, deletions, insertions, and mutations affecting amino acids or splicing sites. For cases with only polymorphic sites, further exon sequencing is needed. This study adds new mutation data to the global HB database, supports research on racial differences in FⅨ gene mutations, and contributes to domestic HB statistics. The results aid in understanding the FⅨ gene\'s role in coagulation, elucidating HB pathogenesis, and providing a basis for future gene therapy.

Adult female and male Gongylonema nematodes were found in the oesophagus of a free-living roe deer (Capreolus capreolus) in Slovenia during passive health surveillance of wildlife. The genus Gongylonema was determined by light microscopy based on the genus-specific cuticular bosses in the anterior part of the parasite. Molecular methods were used to confirm the species Gongylonema pulchrum, which has zoonotic potential. Although Gongylonema species are considered common and distributed worldwide, this is the first report of G. pulchrum in an animal on the territory of Slovenia and the first molecular report in a roe deer worldwide. The parasite is likely to be underdiagnosed, misdiagnosed or goes unnoticed as the animals show little or no clinical signs and minor pathological lesions. Slaughterhouse workers, hunters and veterinarians should be aware of this elusive parasite. Examination and evisceration of the upper digestive tract of animals should therefore be carried out more carefully.

Babesia ovis, transmitted by Rhipicephalus bursa ticks, is the causative agent of ovine babesiosis, a disease characterized by fever, anemia, hemoglobinuria, and high mortality in sheep. This study investigates whether sheep that survived babesiosis without treatment can serve as a source of infection for B. ovis-free host-seeking R. bursa larvae in a later season. Three donor sheep were experimentally infected with B. ovis, and after six months, persistence of B. ovis was assessed through blood and tick transmission experiments. Blood from donor sheep was intravenously injected into three recipient sheep, while donor sheep were also infested with B. ovis-free R. bursa larvae. Engorged nymphs molted to adults, and new recipient sheep were infested with these ticks. All recipient sheep were monitored for B. ovis for 100 days using microscopic, serological, and molecular approaches. The presence of B. ovis was confirmed in the recipient sheep that received blood, leading to clinical infection in two. However, no B. ovis was detected in recipient sheep infested with ticks. These results suggest that sheep recovering from B. ovis infection do not serve as a source of infection for R. bursa larvae in subsequent seasons.

When the polymerase chain reaction (PCR) is used to amplify complex templates such as metagenomic DNA using single or degenerate primers, preferential amplification of templates (PCR bias) leads to a distorted representation of the original templates in the final amplicon pool. This bias can be influenced by mismatches between primers and templates, the locations of mismatches, and the nucleotide pairing of mismatches. Many studies have examined primer-template interactions through interrogation of the final products of PCR amplification with controlled input templates. Direct measurement of primer-template interactions, however, has not been possible, leading to uncertainty when optimizing PCR reactions and degenerate primer pools. In this study, we employed a method developed to reduce PCR bias (i.e., Deconstructed PCR, or DePCR) that also provides empirical data regarding primer-template interactions during the first two cycles of PCR amplification. We systematically examined interactions between primers and templates using synthetic DNA templates and varying primer pools, amplified using standard PCR and DePCR protocols. We observed that in simple primer-template systems, perfect match primer-template interactions are favored, particularly when mismatches are close to the 3\' end of the primer. In more complex primer-template systems that better represent natural samples, mismatch amplifications can dominate, and heavily degenerate primer pools can improve representation of input templates. When employing the DePCR methodology, mismatched primer-template annealing led to amplification of source templates with significantly lower distortion relative to standard PCR. We establish here a quantitative experimental system for interrogating primer-template interactions and demonstrate the efficacy of DePCR for amplification of complex template mixtures with complex primer pools.

Neoadjuvant chemotherapy (NAT) plays a crucial role in breast cancer (BC) treatment, both in advanced BC and in early-stage BC, with different rates of pathological complete response (pCR) among the different BC molecular subtypes. Imaging monitoring is mandatory to evaluate the NAT efficacy. This study evaluates the diagnostic performance of Contrast-Enhanced Mammography (CEM) in BC patients undergoing NAT. This retrospective two-center study included 174 patients. The breast lesions were classified based on the molecular subtypes in hormone receptor (HR+)/HER2-, HER2+, and triple-negative breast cancer (TNBC). The histopathological analysis performed following surgery was used as a reference standard for the pCR. Sensitivity, specificity, PPV, and NPV were measured overall and for the different subtypes. We enrolled 174 patients, 79/174 (46%) HR+/HER2-, 59/174 (33.9%) HER2+, and 35/174 (20.1%) TNBC; the pCR was found in 64/174 (36.8%), of which 57.1% were TNBCs. In the total population, the CEM sensitivity and specificity were 66.2% and 75.2%, with a PPV of 61.4% and an NPV of 78.8%. The highest specificity (80.9%) and NPV (91.7%) were found in HR+/HER2-, while the highest sensitivity (70%) and PPV appeared (73.7%) in TNBC. The results indicate that CEM is a valid tool to assess the pCR, with different performances among the subtypes of BC.