Alteration of ß-glucan in the emerging fungal pathogen Candida auris leads to immune evasion and increased virulence.

Candida auris is an emerging pathogenic yeast that has been categorized as a global public health threat and a critical priority among fungal pathogens. Despite this, the immune response against C. auris infection is still not well understood. Hosts fight Candida infections through the immune system that recognizes pathogen-associated molecular patterns such as ß-glucan, mannan, and chitin on the fungal cell wall. In this study, levels of ß-glucan and mannan exposures in C. auris grown under different physiologically relevant stimuli were quantified by flow cytometry-based analysis. Lactate, hypoxia, and sublethal concentration of fluconazole trigger a decrease in surface ß-glucan while low pH triggers an increase in ß-glucan. There is no inverse pattern between exposure levels of ß-glucan and mannan in the cell wall architecture among the three clades. To determine the effect of cell wall remodeling on the immune response, a phagocytosis assay was performed, followed by quantification of released cytokines by ELISA. Lactate-induced decrease in ß-glucan leads to reduced uptake of C. auris by PMA-differentiated THP-1 and RAW 264.7 macrophages. Furthermore, reduced production of CCL3/MIP-1⍺ but not TNF-⍺ and IL-10 were observed. An in vivo infection analysis using silkworms reveals that a reduction in ß-glucan triggers an increase in the virulence of C. auris. This study demonstrates that ß-glucan alteration occurs in C. auris and serves as an escape mechanism from immune cells leading to increased virulence.

Impact of a spatial repellent intervention on Anopheles kdr insecticide resistance allele in Sumba, Indonesia.

BACKGROUND: The emergence of insecticide resistance and outdoor transmission in malaria-endemic areas underlines the urgent need to develop innovative tools, such as spatial repellents (SR), that may circumvent this residual transmission. With limited options for effective insecticides, regular resistance monitoring is warranted for selecting and using appropriate tools. This study evaluates the pyrethroid knockdown resistance (kdr) allele before and after implementing a transfluthrin-based spatial repellent (SR) intervention in placebo-treated clusters. METHODS: This study looks at the frequency distribution of the kdr allele in Sumba Island from June 2015 to August 2018. Insecticide susceptibility tests were carried out on female Anopheles sp. aged 3-5 days against permethrin 21.5 µg/ml, deltamethrin 12.5 µg/ml, and transfluthrin 10 µg/ml using CDC bottle assay. PCR sequencing of representative samples from adult mosquito collections and insecticide tests revealed the presence of kdr mutations (L1014F and L1014S) in the VGSC gene. RESULTS: A total of 12 Anopheles species, Anopheles tesselatus, Anopheles. aconitus, Anopheles barbirostris, Anopheles kochi, Anopheles annularis, Anopheles maculatus, Anopheles sundaicus, Anopheles flavirostris, Anopheles balabacensis, Anopheles indefinitus, Anopheles subpictus, and Anopheles vagus were analysed. Anopheles vagus and An. sundaicus predominated in the larval populations. Susceptibility assays for all insecticides identified fully susceptible phenotypes in all species examined. Anopheles increasing frequency of kdr mutant alleles during the 3 year SR deployment was observed in both SR-treated and placebo areas, a statistically significant increase occurred in each arm. However, it is unclear how significant SR is in causing the increase in mutant alleles. The L1014S, knockdown resistance east type (kdr-e) allele was detected for the first time among the mosquito samples in this study. The L1014F, knockdown resistance west type (kdr-w) allele and heteroduplex form (wild-type-mutant) were found in almost all Anopheles species examined, including An. vagus, An. aconitus, An. subpictus, An. tesselatus, An. annularis, An. flavirostris and An. sundaicus. CONCLUSION: The presence of fully susceptible phenotypes over time, along with an increase in the frequency distribution of the L1014F/S mutations post-intervention, suggest drivers of resistance external to the study, including pyrethroid use in agriculture and long-lasting insecticidal nets (LLINs). However, this does not negate possible SR impacts that support resistance. More studies that enable the comprehension of possible SR-based drivers of resistance in mosquitoes need to be conducted.

Prediction of the Signaling Pathway in Polycystic Ovary Syndrome Using an Integrated Bioinformatics Approach.

Objectives The purpose of this study was to define the underlying biological mechanisms of PCOS utilizing the protein-protein interaction networks that were constructed based on the putative disease-causing genes for PCOS. Design No animals were used in this research because this study is an In-Silico studies which mainly uses softwares and online analysis tools. Participants/Materials, Settings Genes datasets related with PCOS were obtained from Genecard. Methods The protein-protein interaction networks (PPIN) of PCOS were created using the String Database after genes related with PCOS were obtained from Genecard. After that, we performed an analysis of the hub-gene clusters extracted from the PPIN using the ShinyGO algorithm. In the final step of this research project, functional enrichment analysis was used to investigate the primary biological activities and signaling pathways that were associated with the hub clusters. Results The Genecard database provided the source for the identification of a total of 1072 potential genes related with PCOS. The PPIN that was generated by using the genes that we collected above contained a total of 82 genes and three different types of cluster interaction interactions. In addition, after conducting research on the PPIN with the shinyGO plug-in, 19 of the most important gene clusters were discovered. The primary biological functions that were enriched in the key clusters that were developed were ovarian stereoidogenesis, breast cancer pathway, regulation of lipid and glucose metabolism by AMPK pathway, and ovarian stereoidogenesis. The integrated analysis that was performed in the current study demonstrated that these hub clusters and their connected genes are closely associated to the pathogenesis of PCOS. Limitations Several of the significant genes that were identified in this study, such as ACVR1, SMAD5, BMP6, SMAD3, SMAD4 and AMH. It is necessary to do additional research using large samples, several centers, and multiple ethnicities in order to verify these findings. Conclusions The integrated analysis that was performed in the current study demonstrated that these hub clusters and their connected genes are closely associated to the pathogenesis of PCOS. This information may possibly bring unique insights for the treatment of PCOS as well as the investigation of its underlying pathogenic mechanism.

The origin and possible mechanism of embryonic cell-free DNA release in spent embryo culture media: a review.

The presence of cell-free DNA in spent embryo culture media (SECM) has unveiled its possible utilization for embryonic ploidy determination, opening new frontiers for the development of a non-invasive pre-implantation genetic screening technique. While a growing number of studies have shown a high concordance between genetic screening using cell-free DNA (cfDNA) and trophectoderm (TE), the mechanism pertaining to the release of cfDNA in SECM is largely unknown. This review aims to evaluate research evidence on the origin and possible mechanisms for the liberations of embryonic DNA in SECM, including findings on the self-correction abilities of embryos which might contribute to the presence of cfDNA. Several databases including EMBASE, PUBMED, and SCOPUS were used to retrieve original articles, reviews, and opinion papers. The keywords used for the search were related to the origins and release mechanism of cfDNA. cfDNA in SECM originates from embryonic cells and, at some levels, non-embryonic cells such as maternal DNA and exogenous foreign DNA. The apoptotic pathway has been demonstrated to eliminate aneuploid cells in developing mosaic embryos which might culminate to the release of cfDNA in SECM. Nonetheless, there is a recognized need for exploring other pathways such as cross-talk molecules called extracellular vesicles (EVs) made of small, round bi-layer membranes. During in vitro development, embryos physiologically and actively expel EVs containing not only protein and microRNA but also embryonic DNA, hence, potentially releasing cfDNA of embryonic origin into SECM through EVs.

The effects of luteinizing hormone as a suppression factor for apoptosis in bovine luteal cells in vitro.

The fate of the corpus luteum, a transient endocrine gland formed and degraded during an oestrous cycle, is decided by various physiological factors, such as luteinizing hormone (LH). As a stimulator of progesterone, LH is known to maintain corpus luteum functional and structural integrity by inhibiting apoptosis, a programmed cell death. Therefore, we aim to investigate its action during the mid-luteal phase hypothesized that LH suppresses the death mechanism of bovine luteal steroidogenic cells (LSC) by analysing the expression of proteins involved. Cultured bovine LSC obtained from corpus luteum were treated for 24 hr with recombinant TNF and IFNG in the presence or absence of LH. The result showed that LH proved to have a protective effect by increased cell viability (p < .05) and prevented DNA fragmentation (p < .05), as demonstrated by the WST-1 colorimetric assay and TUNEL assay. Expression analysis of mRNA and protein levels showed that LH altered the expression of BCL2 (p < .05), CASP3 (p < .05), FAS (p < .05), and BAX (p < .05) to support cell survival. In conclusion, our study suggests that LH prolongs the corpus luteum life span through the anti-apoptotic mechanism by increasing cell viability and suppressing apoptosis-related genes and protein expression.

Identification and functional characterization of Penicillium marneffei major facilitator superfamily (MFS) transporters.

PENICILLIUM MARNEFFEI: is a thermally dimorphic fungus that causes penicilliosis, and become the third-most-common opportunistic fungal infection in immunocompromised patients in Southeast Asia. Azoles and amphotericin B have been introduced for the treatment, however, it is important to investigate possible mechanisms of azole resistance for future treatment failure. We identified 177 putative MFS transporters and classified into 17 subfamilies. Among those, members of the Drug:H+ antiporter 1 subfamily are known to confer resistance to antifungals. Out of 39 paralogs, three (encoded by PmMDR1, PmMDR2, and PmMDR3) were heterologously overexpressed in S. cerevisiae AD∆ conferred resistance to various drugs and compounds including azoles, albeit to different degrees. PmMDR1-expressing strain showed resistance to the broadest range of drugs, followed by the PmMDR3, and PmMDR2 conferred weak resistance to a limited range of drugs. We conclude that PmMDR1 and PmMDR3, may be able to serve as multidrug efflux pumps.

Non-invasive pre-implantation genetic testing's reliability for aneuploidy using Cell-free DNA in embryo culture media.

OBJECTIVE: The presence of embryonic cell-free DNA (cfDNA) in spent embryo culture media (SECM) may offer valuable advantages for non-invasive testing of embryo ploidy or genetic characteristics compared to trophectoderm (TE) biopsy. This study aimed to assess the diagnostic potential of SECM cfDNA as a non-invasive sample for chromosomal copy number testing in blastocysts within the clinical setting of in-vitro fertilization. METHOD: This prospective observational study collected 28 SECM cfDNA samples matched with TE biopsy samples from 21 infertile couples who underwent IVF-PGT-A cycles. SECM samples were obtained from blastocysts that were cultured for approximately 5/6 days in an uninterrupted time-lapse incubator. Both sets of samples were collected during the biopsy procedure. The Variseq Illumina platform was utilized for ploidy measurement. The study evaluated the informativity and interpretability of SECM cfDNA, concordance of general ploidy status, and sex chromosome agreement between the two sample types. RESULTS: SECM cfDNA had a high informativity rate (100 %) after double amplification procedure, with a result interpretability of 93 %. Two out of the 28 SECM cfDNA samples were uninterpretable and regarded as overall noise samples. The diagnostic potential of SECM cfDNA, when compared to TE biopsy the standard reference, was relatively low at 50 %. Maternal DNA contamination remains the major obstacle that hinders the widespread clinical adoption of SECM cfDNA in the routine practice of pre-implantation genetic testing for aneuploidy within IVF settings. CONCLUSION: A significant modification must be implemented in the IVF laboratory to minimize DNA contamination and this necessitates suggesting adjustments to oocyte denudation, embryo culture media preparation, and sample collection procedures.

IL-6 and IL-10 Levels in Rats Blood Plasma as Immune Responses Post Radioiodine (I-131) Administration.

OBJECTIVE: The purpose of this study was to analyze the effect of oral administration of radioiodine (I-131) on the immune responses (interleukin 6 and 10) as biodosimetry markers and to support clinical trials of I-131 solution. METHODS: The design of this study was an in vivo experimental study using twenty-seven male rats (Rattus norvegicus strain Sprague-Dawley) given 100 µL of I-131 solution at a dose of 260 µCi. Blood plasma was taken at 0.25, 0.5, 1, 3, 24, 48, 120, and 168 hours post oral I-131 administration, respectively. Rats without radioiodine administration as a control group. The levels of IL-6 and IL-10 were measured using the enzyme-linked immunosorbent assay (ELISA) method. Statistical analysis was carried out with one-way ANOVA using SPSS version 25 software. RESULT: IL-6 level began to significantly increase at 0.25 hours post administration of I-131 (14.4 pg/mL ± 2.52 pg/mL, p=0.02). During 7 days of observation, IL-6 levels had 2 peaks of highly significant increase at 0.5 hours (43.57 ± 5.28, p<0.001) and 120 hours (24.08 ± 2.69, p<0.001 compared to control (5.44 ± 0.95 pg/mL). IL-10 level began to significantly increase at 0.25 hours (30.32 ± 3.22 pg/mL, p=0.03) compared to controls (20.61 ± 1.59 pg/mL). CONCLUSION: The highest increase in IL-6 and IL-10 levels occurred respectively in the first 0.5 hours 8 times and in the first 0.25 hours 1.47 times compared to controls. Internal irradiation with radioiodine resulted in a significant increase in immune cells in exposed blood plasma characterized by the production of the cytokines IL-6 and IL-10. This appears to be a response of immune cells to reduce or stop inflammatory reactions through the release of anti-inflammatory cytokines in an effort to prevent excessive inflammatory responses that can damage cells and tissues.

Luteoprotective roles of luteinizing hormone are mediated by not only progesterone production but also glucocorticoid conversion in bovine corpus luteum.

Luteinizing hormone (LH) is known as a key regulator of corpus luteum (CL) function, but the luteoprotective mechanisms of LH in the maintenance of bovine CL function are not well understood. The current study investigated if LH increases cell viability and induces cortisol conversion, and if the luteoprotective action of LH is mediated by stimulating the local production and action of progesterone (P4) and/or cortisol. Cultured bovine luteal cells obtained at the mid-luteal stage (Days 8-12 of the estrous cycle) were treated for 24 hr with LH (10 ng/ml) with/without onapristone (OP, a specific P4 receptor antagonist; 100 µM), cortisone (1 µM), and aminoglutethimide (AGT, a specific inhibitor of cytochrome P450 side-chain cleavage; 100 µM). LH with and without OP significantly increased the mRNA and protein expressions of 11ß-hydroxysteroid dehydrogenase (HSD11B) 1, but did not affect the mRNA or protein expression of HSD11B2. These treatments also significantly increased HSD11B1 activity. Cell viability was significantly increased by LH alone or by LH in combination with cortisone and OP. LH in combination with OP or AGT significantly decreased cell viability as compared to LH alone. The overall results suggest that LH stimulates not only P4 production but also HSD11B1 expression, thereby increasing the cortisol concentration in the bovine CL, and that LH prevents cell death through these survival pathways. LH may consequently support CL function during the luteal phase in cattle.

Luteoprotective mechanisms of prostaglandin F2α stimulated by luteinizing hormone in the bovine corpus luteum.

Luteinizing hormone (LH) regulates several ovarian functions. However, the luteoprotective mechanisms of LH involved in the maintenance of bovine corpus luteum (CL) function are not well understood. Since prostaglandin F2α (PGF), PGE2 and progesterone (P4) are well documented as antiapoptotic factors in the bovine CL, we hypothesized that LH protects the CL by stimulating the local production and action of PGF, PGE2 and P4. Cultured bovine luteal cells obtained at the mid-luteal stage (days 8-12 of the estrous cycle) were treated with LH (10 ng/ml), onapristone (OP: a specific P4 receptor antagonist, 100 µM) and indomethacin [INDO; a cyclooxygenase (COX) inhibitor, 100 µM] for 24 h. LH with and without OP significantly increased the mRNA and protein expressions of COX-2, PGF synthase and carbonyl reductase (P<0.05) but not the mRNA and protein expressions of COX-1 and PGE synthase in bovine luteal cells. In addition, these treatments significantly increased PGF and P4 production (P<0.05) but not PGE2 production. Luteal cell viability was significantly increased by LH alone (P<0.05), but LH-increased cell viability was reduced by LH in combination with INDO as well as OP (P<0.05). The overall results suggest that LH prevents luteal cell death by stimulating luteal PGF and P4 production and supports CL function during the luteal phase in cattle.

microRNAs as A Biomarker to Predict Embryo Quality Assessment in In Vitro Fertilization.

Embryo selection for in vitro fertilization (IVF) is an effort to increase the success rate of embryo implantation. Factors influencing the success of embryo implantation include embryo quality, endometrial receptivity, embryo characteristics, and maternal interactions. Some molecules have been found to influence these factors, but their regulatory mechanisms are unclear. MicroRNAs (miRNAs) are reported to play an essential role in the embryo implantation process. miRNAs are small non-coding RNAs consisting of only 20 nucleotides that play an essential role in the stability of gene expression regulation. Previous studies have reported that miRNAs have many roles and are released by cells into the extracellular environment for intracellular communication. In addition, miRNAs can provide information related to physiological and pathological conditions. These findings encourage research development in determining the quality of embryos in IVF to increase the implantation success rate. Moreover, miRNAs can provide an overview of embryo-maternal communication and potentially be noninvasive biological markers of embryo quality, which could increase assessment accuracy while reducing mechanical damage to the embryo itself. This review article summarizes the involvement of extracellular miRNAs and the potential applications of miRNAs in IVF.

Ultrastructure of Javaen barb fish Systomus orphoides Valenciennes, 1842 spermatozoa (Cypriniformes: Cyprinidae) by electron microscopes.

Javaen barb fish Systomus orphoides Valenciennes, 1842 (Cypriniformes: Cyprinidae) is a freshwater fish whose population is declining and threatened with extinction. In this study, the ultrastructure of spermatozoa of Javaen barb fish (S. orphoides) was studied using transmission and scanning electron microscopy. The spermatozoa of S. orphoides are relatively simple cells composed of a spherical head, a short midpiece, and a flagellum, as in most Cyprinidae. The ultrastructure is characterized by the absence of acrosome, the total length of spermatozoa is 27.16 ± 4.5 µm, and the head has spherical with a length of 1.84 ± 0.10 µm and width of 1.55 ± 0.15 µm containing a nucleus, midpiece region containing the proximal and distal centrioles and mitochondria. Two or three mitochondria surrounding the axoneme (with a 9 + 2 microtubular pattern). Ultrastructural analyses by SEM and TEM of Javaen barb fish spermatozoa cells are very consistent with those of Cyprinidae. This study provides the ultrastructure information of S. orphoides spermatozoa in the Cyprinidae family this research could be useful in increasing reproductive efficiency and further prevent the extinction of this species.

The Fungal and Bacterial Interface in the Respiratory Mycobiome with a Focus on Aspergillus spp.

The heterogeneity of the lung microbiome and its alteration are prevalently seen among chronic lung diseases patients. However, studies to date have primarily focused on the bacterial microbiome in the lung rather than fungal composition, which might play an essential role in the mechanisms of several chronic lung diseases. It is now well established that Aspergillus spp. colonies may induce various unfavorable inflammatory responses. Furthermore, bacterial microbiomes such as Pseudomonas aeruginosa provide several mechanisms that inhibit or stimulate Aspergillus spp. life cycles. In this review, we highlighted fungal and bacterial microbiome interactions in the respiratory tract, with a focus on Aspergillus spp.

Evolution and immunopathology of chikungunya virus informs therapeutic development.

Chikungunya virus (CHIKV), a mosquito-borne alphavirus, is an emerging global threat identified in more than 60 countries across continents. The risk of CHIKV transmission is rising due to increased global interactions, year-round presence of mosquito vectors, and the ability of CHIKV to produce high host viral loads and undergo mutation. Although CHIKV disease is rarely fatal, it can progress to a chronic stage, during which patients experience severe debilitating arthritis that can last from several weeks to months or years. At present, there are no licensed vaccines or antiviral drugs for CHIKV disease, and treatment is primarily symptomatic. This Review provides an overview of CHIKV pathogenesis and explores the available therapeutic options and the most recent advances in novel therapeutic strategies against CHIKV infections.

The Effect of Follitropin Alfa in Controlled Ovarian Stimulation Protocol for In Vitro Fertilization Cycles.

Background: Follitropin alfa (FA) is one of the most widely used exogenous gonadotropins in both agonist and antagonist protocols for controlled ovarian stimulation (COS) and in vitro fertilization (IVF). However, reports of its effectiveness are limited, particularly in terms of its impact on overall IVF outcomes and ovarian hyperstimulation syndrome (OHSS). Therefore, in this study, FA competency was investigated by evaluating its effect on IVF outcomes and OHSS, administering agonist and antagonist COS protocols. Methods: A retrospective study with 120 subjects was conducted. Outcomes comprising the number of retrieved and fertilized oocytes, quality of embryos, and clinical pregnancies were assessed. Statistical correlation between FA dose, IVF outcomes, and the incidence of OHSS was also analyzed. All statistical analyses were performed at 95% confidence level. Results: There was no significant difference in both protocols regarding retrieved oocytes (p=0.604), fertilized oocytes (p=0.761), embryo quality including good, average, poor embryo (p=0.875, p=0.565, p=0.785), and clinical pregnancy (p= 0.844). However, FA doses in the agonist protocol were shown notably higher (p= 0.001). Negative correlations were also observed between FA dose and the number of retrieved oocytes (r=-0.255, p<0.01), fertilized oocytes (r=-0.296, p<0.01), and good quality embryos (r=-0.231, p<0.05). Conclusion: Our study suggested that FA yields similar outcomes in both COS protocols, but agonist protocols require higher doses of FA and evaluation of its effect on OHSS is an important area of research for further investigation.

Anti-Müllerian hormone independently affect mtDNA copy number in human granulosa cells.

BACKGROUND: Recently, as a delayed childbearing trend is emerging in modern women's adulthood, diminished reproductive potential due to age-related changes is more prevalent. Reduction in the abundance of mitochondrial DNA (mtDNA) copies and circulating anti-Müllerian hormone (AMH) have been separately reported with aging, contributing to the decrease in successful reproduction. However, there are limited reports on the impact of age on mtDNA and AMH in the same individual and whether mtDNA copy numbers are influenced by age and AMH. METHODS: In the present study, we utilized a real-time quantitative PCR (RT-qPCR) to quantify the mtDNA copy number of granulosa cells obtained from 43 women undergoing an in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) program. RESULTS: According to our analysis, a significant correlation was observed between age and mtDNA copy number (r = -0.54, P < 0.001) and between age and AMH level (r = -0.48, P < 0.001) of the same individual. There was also a positive correlation between mtDNA copy number and AMH (r = 0.88, P < 0.001) with AMH level falling as mtDNA decreases. In our regression, age and AMH were shown to have low collinearity (VIF = 1.297) but only AMH was correlated with mtDNA quantity (P < 0.001). CONCLUSION: Our study suggests that both mtDNA and AMH abundance are influenced by age and that AMH levels independently affect mtDNA copy number regardless of age. Further research is required to understand the role of AMH on mitochondria bioenergetics.

Low-cost SYBR Green-based RT-qPCR assay for detecting SARS-CoV-2 in an Indonesian setting using WHO-recommended primers.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent for the ongoing coronavirus disease 2019 (COVID-19) pandemic. For laboratory diagnosis, low-cost detection of SARS-CoV-2 is urgently needed, particularly in developing countries with limited resources. Probe- or TaqMan-based real-time reverse transcription polymerase chain reaction (RT-qPCR) is currently the gold standard for diagnosing infected individuals, as recommended by the World Health Organization (WHO). However, this assay is expensive, making it difficult to use for diagnosis on a large scale. Therefore, in this study, we develop and validate an alternative approach for RT-qPCR diagnosis by employing the DNA intercalating dye SYBR Green. We evaluate and use two WHO-recommended primers, namely CCDC-N and HKU-ORF1b-nsp14. The compatibility of the two primers was tested in silico with Indonesian SARS-CoV-2 genome sequences retrieved from the GISAID database and using bioinformatic tools. Using in vitro-transcribed RNA, optimization, sensitivity, and linearity of the two assays targeting the N and Nsp-14 genes were carried out. For further evaluation, we used clinical samples from patients and performed the SYBR Green-based RT-qPCR assay protocol in parallel with TaqMan-based commercial assay. Our results show that our methodology performs similarly to the broadly used TaqMan-based detection method in terms of specificity and sensitivity and thus offers an alternative assay for the detection of SARS-CoV-2 RNA for diagnostic purposes.

Unravelling the Molecular Identification and Antifungal Susceptibility Profiles of Aspergillus spp. Isolated from Chronic Pulmonary Aspergillosis Patients in Jakarta, Indonesia: The Emergence of Cryptic Species.

Cryptic species of Aspergillus have rapidly increased in the last few decades. Chronic pulmonary aspergillosis (CPA) is a debilitating fungal infection frequently affecting patients with previous TB. The identification and antifungal susceptibility profiles of different species of Aspergillus are important to support the management of CPA. The aim of this study was to describe the molecular and susceptibility profiles of Aspergillus isolated from CPA patients. The species identity of isolates was determined by combined DNA analyses of internal transcribed space (ITS), partial ß-tubulin genes, and part of the calmodulin gene. We revealed a high (27%) prevalence of cryptic species among previous tuberculosis patients with persistent symptoms. Twenty-nine (49%) patients met the criteria for diagnosis of CPA with 24% containing Aspergillus cryptic species. This is the first report of five cryptic Aspergillus species from clinical isolates in Indonesia: A. aculea tus, A. neoniger, A. brunneoviolacues, A. welwitschiae, and A. tubingensis. Significantly, there was decreased sensitivity against itraconazole in the CPA group (66% susceptible to itraconazole) compared to the non-CPA group (90% susceptible to itraconazole) (p = 0.003). The species-level characterisation of Aspergillus and its antifungal susceptibility tests demands greater attention to better the management of CPA patients.

Effect of egg yolk of free-range chicken and methanol as a cryoprotective agent for the sperm preservation of cyprinid fish, Neolissochilus soroides (Valenciennes, 1842).

The objective of this study was to determine the optimum concentration of egg yolk of free-range chicken as a cryoprotective agent on cyprinid fish, Neolissochilus soroides sperm after 48 h frozen. One level of methanol (10%) combined with six levels of egg yolk solution (0%, 5%, 10%, 15%, 20% and 25%) were tested. Fish Ringer's solution was used as an extender. The diluted sperm was equilibrated for 10 min at 5 °C, then kept at -10 °C temperature for 48 h. Sperm was thawed for 1 min at 40 °C. Spermatozoa viability, abnormality, and fertilization rates were analysed afterwards. The one-way ANOVA showed that the combination methanol with several concentrations of egg yolk solution had a significant effect on spermatozoa viability, abnormality, and fertilization rates (P < 0.05) by improving semen character. The study revealed that the 5% egg yolk solution combined with 10% methanol resulted in the highest rates of viability (82.13 ± 1.75%) and fertility rates (92.96 ± 1.94%), with the lowest abnormality (25.25 ± 2.22%). A 5% egg yolk solution was identified as the best cryoprotective agent for N. soroides spermatozoa preservation at -10 °C for 48 h.

Analysis of Bio-Risk Management System Implementation in Indonesian Higher Education Laboratory.

Developing countries face various challenges in implementing bio-risk management systems in the laboratory. In addition, educational settings are considered as workplaces with biohazard risks. Every activity in a laboratory facility carries many potential hazards that can impact human health and the environment and may cause laboratory incidents, including Laboratory Acquired Infections (LAIs). In an effort to minimize the impact and occurrence of these incidents, it is necessary to evaluate the implementation of a bio-risk management system in every activity that involves handling biological agents. This study was conducted in an Indonesian higher-education institution, herein coded as University Y. This is a descriptive, semi-quantitative study aimed at analysing and evaluating the implementation of the bio-risk management systems used in laboratories by analysing the achievements obtained by each laboratory. The study used primary data that were collected using a checklist which referred to ISO 35001:2019 on Laboratory Bio-risk Management. The checklist consisted of 202 items forming seven main elements. In addition, secondary data obtained from literature and document review were also used. The results show that out of 11 laboratories examined, only 2 laboratories met 50% of the requirements, which were Laboratory A and B, achieving good performance. Regarding the clauses of standards, a gap analysis identified leadership, performance evaluation, and support as elements with the lowest achievement. Therefore, corrective action should be developed by enhancing the commitment from management as well as improving documentation, policy, education and training.