Estrogen has a protective role on auditory function. It may have an excitatory action on auditory nerve fibers and can have a neuroprotective effect. Progesterone has a mainly inhibitory action on the central nervous system, which may balance the mainly excitatory action of estrogen. To determine changes in hearing performance with pure tone audiometry (PTA), tympanometry, distortion product otoacoustic emissions (DPOAEs), and auditory brainstem responses (ABR) as hormonal changes occur from follicular to luteal phase. Twenty healthy female volunteers (age 19 ± 30 years) with normal menstrual cycle and without any hearing problems are included in this case-control study. Hearing evaluation was performed on the 13th day of the menstrual cycle (follicular phase) and then on the 22nd day . All of the participants had normal results in follicular phase. In luteal phase, four cases showed abnormalities as follows: reduced hearing thresholds 250 Hz (mean= 15 dBHL), increased amplitudes of DPOAE (mean= 3 dBspl), decreased middle ear pressure (mean= -110 dapa), and delayed ABR interpeak latencies (mean of IPLs I-III= 0.4 and mean of IPLs III-V= 0.6 ms). In some women, changing of ovarian hormones may induce fluctuating hearing and increased progesterone in luteal phase can lead to abnormal outcomes in auditory function. However, elevated estrogen modifies its consequences in follicular phase. Estrogen and progesterone are female sex hormones and their secretion change during menstruation. Estrogen (oestrogen) rises during the discharge of the white body or follicular phase , 2. It iEstrogen has a protective role on auditory function. It may have an excitatory action on auditory nerve fibers and can have a neuroprotective effect. At the cochlear level, estrogen receptors alpha and beta have been identified in the inner ear, including spiral ganglion type I cells, the stria vascularis, and cochlear blood vessels. Estrogen may influence auditory transmission, while the receptors in the stria vascularis may affect fluid electrolyte balance in the cochlear fluids and modulating cochlear blood flow .Reduction of estrogen results in poor performance of the auditory pathway. Blocking estrogen receptors alpha and beta had led to a decline in suppression of DPOAEs as a result of a reduction of the medial olivocochlear suppression. This decline could be similar to what occurs with aging and precedes the onset of age related hearing loss, which may suggest that estrogen has a protective role on auditory function, in conjunction with its neuroprotective effect. It has been observed that the decrease in estrogen levels, e.g., after menopause or in Turner's syndrome, is associated with an increased frequency of neurodegenerative disorders. Another proposed mechanism would implicate estrogen as a free radical scavenging antioxidant .Important point is that the entire central nervous system, auditory, and vestibular structures are dimorphism; i.e., there is a gender difference in serotonergic system. It has been reported that there is an increase in 5-HT activity in the female brain compared with the male and a decrease in the whole brain 5-HT synthesis in women compared with men. It was also found to control the gain the inferior colliculus neural responses in a positive or negative direction depending on the type of auditory stimuli .Progesterone is also secreted by the adrenal glands of the central nervous system. It increases in luteal phases of the menstrual cycle, pregnancy, and embryogenesis function. It has a variety of important functions in the body and is also a crucial metabolic intermediate in the production of other endogenous steroids, including the sex hormones and corticosteroids, and plays an important role in brain function as a neurosteroid , 5.Progesterone has a mainly inhibitory action on the central nervous system, which may balance the mainly excitatory action of estrogen. It was found to decrease 5-HT levels and this may affect auditory processing indirectly and may cross-react with other steroid receptors  present in the cochlea or more proximal areas of the auditory system. It may also influence the auditory system through its interaction with the steroid binding sites on GABA-A receptors acting as a GABA-A agonist, which are present throughout the auditory system . ChangesSo, this study aimed to determine changes in hearing performance with pure tone audiometry (PTA), tympanometry, distortion product otoacoustic emissions (DPOAEs), and auditory brainstem responses (ABR) as hormonal changes occur from follicular to luteal phase.The contract for this project was signed on July 22, 2014, with number 9304312272. The sample size consisted of twenty healthy female of age 19–30 years. They studied from July 22, 2014, to July 15, 2015.Study was case-control; each one was compared with herself and subjected to a two-stage assessment of hearing function by standard clinical audiological battery tests. Assessments started on the 13th day of the menstrual cycle (follicular phase) and then on the 22nd day . The levels of estradiol and progesterone were measured. We chose the people who were sure to be healthy at the first stage of the evaluation. The results of the first assessment phase were recorded as the baseline (control group). In the second stage , changes  were recorded as case group and compared to the first stage. Ovulation was estimated by measuring basal body temperature (BBT). Subjects were instructed to note down their menstrual calendar for at least 2 months along with their BBT for detecting the day of ovulation. The study was conducted under standard laboratory conditions (room temperature 26 ± 2°C) in sound proof room. This study compared PTA, tympanometry, and DPOAE and ABR between pre- and postovulatory phases of menstrual cycle using Madsen: OB-822, Maico: MI.34, Homoth, and Labat Epic-plus.2, menstrual cycle = 28 days, level of estradiol for the 13th day in the follicular phase > 600 pg/ml and for the 22nd day in the luteal phase < 250 pg/ml, and level of progesterone for similar days < 50 and > 2000 ng/dl, respectively.Inclusion criteria are as follows: hearing status = normal, gynecological history = normal, body mass index (BMI) = 18.5 to 24.9 kg/mExclusion criteria are as follows: history of any hearing loss, genital infections, ovarian failure, Turner syndrome, hypothyroidism, Stein-Leutal syndrome, menopause, pregnant or lactating mother, polycystic ovary syndrome, early puberty, receiving steroid hormones, therapy for depression, having history of head injury, stroke, heart attack, disorders of endocrine function, metabolic neoplastic pathologies, anorexia nervosa, liver cellular necrosis, harmful doses of nicotine or alcohol, and alcoholic liver. This information was obtained based on medical records and case history.Procedure: normal PTA thresholds were in the frequency range of 250–8000 Hz of less than 15 dB  , it is an absent response. What is sufficient SNR? This depends on noise floor calculations used by the instrumentation. Some instrumentation calculates the mean noise floor with response variability (standard deviations) incorporated into the measurement. If this is the case, a 3–4 dB SNR is sufficient to detect a response). The f 1/ f 2 ratio was fixed at 1.22 and stimulus levels were held constant at L1; 65 dBspl, and L2; 55 dBspl. The 2f1−f2 DPOAE amplitudes were recorded at frequencies 1.0, 1.5, 2.0, 3.0, 4.0, 6.0, and 8.0 KHz [In the first stage of the evaluation, it was important for us to select people whose middle ear pressure has a minimum deviation of zero dopa. Normal DPOAE amplitudes were 3 dBspl above the noise floor  during the luteal phase complained of a feeling of fullness in the ears, heaviness of the head, and nasal congestion, but they did not have sensation of dizziness or vertigo. All of these seven cases had abnormal results in eustachian tube function test {. Our results showed that, in some sensitive women, changes in ovarian hormones can lead to abnormal outcomes in auditory tests. The cause of this problem seems to be due to individual differences. It seems that the auditory system in some women is more sensitive to hormone changes. In this group of women, increasing the level of progesterone in luteal phase may partially affect the function of the auditory system.Our results are consistent with the research of other researchers. It may have a negative effect on both peripheral (DPOAE) and central auditory function and support the positive role of estrogen on hearing function. Also, increase in interstitial fluids (possibly due to progesterone) affects the eustachian tube function and thus may lead to poorer thresholds , 11.Estrogen plays a main role in maintaining the normal hearing of young and middle-aged women in comparison to postmenopausal females , 11, 12.Because of the excitatory and neuroprotective role of estrogen, the high tone hearing thresholds in males compared to females are poorer and due to inhibitory role of progesterone, hearing thresholds for low frequencies are weaker in women than their male counterparts . In the Also, the shorter ABR latencies during the periovulatory phase of the cycle may suggest that the high estrogen level is associated with shorter ABR latencies. The higher level of estrogen may alter the speed of sensory neurotransmission in the brain stem by modulating glutamate transmission. The higher levels of estrogen may result in an increase in neurosteroids  which facilitate GABA inhibition in the auditory midbrain. The difference between the ABR latencies in the periovulatory phase and luteal phase would be compatible with progesterone blunting the effect of estrogen, by inhibiting its action in the auditory brainstem . SeveralIncreasing levels of estrogen rises nerve steroids, which facilitates gamma aminobutyric acid (GABA) inhibition in the brain. Estrogen therapy on a species of mice increased the level of allopregnanolone in the central and peripheral level and the difference in latency of the pre- and postmenopausal women with ovarian hormonal changes and the stimulatory and inhibitory effects of estrogen and progesterone can be argued . ChangesReduction of DPOAE amplitudes following age increases in various species of motility and after menopause. Direct hearing loss due to age is confirmed by pure audiometry and menopause . The lowStudies show that women with premenstrual syndrome (PMS) have had irregular responses in the presence of stress compared to noninfected women. These changes in PMS women have a potential impact on auditory function . ChangesContraceptive drugs inhibit the levels of estrogenic hormone levels. In women, who have contraceptive medications compared to women who have not, the range of OAEs is smaller and the responses are weaker and similar to men , 20.The physiological changes in pregnancy may affect auditory function mimicking the auditory dysfunction seen in Meniere's disease, i.e., low frequency elevated thresholds, with lowering of uncomfortable loudness levels. The higher levels of ovarian hormones in pregnancy may shorten ABR wave latencies, but the higher level of progesterone may blunt this effect. Neural conduction in the brain stem may be slower during pregnancy due to the fact that higher levels of steroids that facilitate GABA inhibition the absolute wave latencies were slightly shorter but did not reach significance in eight pregnant women in the third trimester compared with age-matched nonpregnant women. However, the I–III, III–V, and I–V interpeak latencies which indicate neural conduction were significantly longer in pregnant women .The auditory system seems to be sensitive to changing female hormones in some women and that is important factor, which can be the reason for reinforcement-attenuation of responses. It can affect the latencies of auditory brainstem responses, the amplitudes of distortion product otoacoustic emissions, the pressure of middle ear, the function of eustachian tube, and low frequency hearing thresholds.


Telocytes (TCs) are newly identified interstitial cells that participate in tissue protection and repair. The present study investigated the mechanisms underlying the protective effect of TCs in a mouse model of respiratory distress.The mouse model of acute respiratory distress syndrome (ARDS) was established by intratracheal instillation of lipopolysaccharide (LPS). After instillation of TCs culture medium, lung injury was assessed, and angiogenesis markers, including CD31 and endothelial nitric oxide synthase (eNOS), were detected by immunofluorescence. Bioinformatics analysis was used to screen significantly differentially expressed microRNAs (miRNAs) in cultured TCs stimulated with LPS, and the regulation of downstream angiogenesis genes by these miRNAs was analysed and verified. PI3K subunits and pathways were evaluated by using a PI3K p110α inhibitor to study the involved mechanisms.In ARDS mice, instillation of TCs culture medium ameliorated LPS-induced inflammation and lung injury and increased the protein levels of CD31 and eNOS in the injured lungs. A total of 7 miRNAs and 1899 mRNAs were differentially regulated in TCs stimulated with LPS. Functional prediction analysis showed that the differentially expressed mRNAs were enriched in angiogenesis-related processes, which were highly correlated with miR-21a-3p. Culture medium from TCs with miR-21a-3p inhibition failed to promote angiogenesis in mouse models of LPS-induced ARDS. In cultured TCs, LPS stimulation upregulated the expression of miR-21a-3p, which further targeted the transcription factor E2F8 and decreased Notch2 protein expression. TCs culture medium enhanced hemangioendothelioma endothelial cells (EOMA cells) proliferation, which was blocked by the miR-21a-3p inhibitor. The PI3K p110α inhibitor decreased vascular endothelial growth factor levels in LPS-stimulated TCs and reversed the enhancing effect of TCs culture medium on EOMA cells proliferation.TCs exerted protective effects under inflammatory conditions by promoting angiogenesis via miR-21a-3p. The PI3K p110α subunit and transcriptional factor E2F8 could be involved in this process. Acute respiratory distress syndrome (ARDS) is a clinical syndrome characterised by acute progression of respiratory failure. According to an international multi-centre research, the prevalence of ARDS was 10.4% of ICU admissions . TherefoTelocytes (TCs) are newly identified mesenchymal cells that play a role in providing nutrition to surrounding cells by cell–cell communication and have post-injury repair and regeneration functions –7. TCs cPI3KCG, a gene encoding a p110γ subunit, which has a protective effect on hypoxic-reoxygenated cardiomyocytes mediated by activation of the PI3K/AKT signalling pathway and inhibition of apoptosis  overnight at 4 °C. Protein expression levels were normalised to those of GAPDH with ImageJ .EOMA cells proliferation was assessed with a colorimetric assay—Cell Counting Kit-8 —following the manufacturer’s protocol. Approximately 4000 EOMA cells/well were seeded in a 96-well plate. After adhesion, EOMA cells were incubated for 24 h with culture medium from TCs transfected with the miR-21a-3p inhibitor or NC in the presence of LPS.The pGL3 reporter vector  was used to generate the plasmids pGL3-WT-E2F8-3′-UTR and pGL3-Mut-E2F8-3′-UTR. Human embryonic kidney cells were co-transfected with pGL3-E2F8-3′-UTR (WT or Mut) and the miR-21a-3p mimic or NC with Lipofectamine 2000 reagent . After incubation for 24 h, luciferase activity was assessed by the Dual-Luciferase Reporter Assay System  according to the manufacturer’s protocol.The concentration of VEGF in the TCs culture medium was measured by a commercial VEGF ELISA kit  according to the manufacturer’s protocol.Live observation of EOMA cells was performed with a Cell-IQ cell culture platform  equipped with a phase contrast microscope (Nikon CFI Achromat phase contrast objective with 10 magnification) and a camera . The equipment was controlled by Imagen software (Chip-Man Technologies). Each group contained 16 replicates of visual fields. Images were acquired at 1-h intervals for 48 h.2O2 for 30 min and heating to boiling in a microwave in citrate buffer for 10 min. After blocking with 5% goat serum in Tris-buffered saline, sections were incubated with diluted primary antibodies  overnight at 4 °C and then with secondary antibodies and 4′,6-diamidino-2-phenylindole (DAPI), separately.Lung tissues were fixed with 10% formalin solution and embedded in paraffin. Each tissue was sectioned at 5 μm and stained with haematoxylin–eosin  according to the manufacturer’s protocol. For immunofluorescence staining, an antigen retrieval protocol was carried out with incubation in 0.3% HData are expressed as the means ± SDs and were analysed by one-way analysis of variance (ANOVA) and Tukey’s multiple comparisons test. A P-value of < 0.05 was considered statistically significant. All statistical analyses were performed with GraphPad Prism 7.04 .The ability for TCs protection was first estimated in ARDS mouse models. LPS stimulation caused inflammatory infiltration, alveolar wall widening, and vessel destruction Fig. a. The prAs angiogenesis is essential in tissue repair, the induction of angiogenic factors in TCs after stimulation with LPS was assessed. In ARDS mice, the expression of the angiogenesis-related marker CD31 and eNOS was downregulated. However, an increase in CD31 and eNOS expression was observed in the WT TCs treatment group but not in the group treated with medium from TCs with miR-21a-3p inhibition  was downregulated with an absolute fold change > 2  and induce angiogenesis by directly targeting PTEN , 31. MorThe transcription factors E2F7/8 were reported to regulate vessel branching via DLL4-Notch approaches  or HIF-1PI3K, especially the Class I catalytic isoforms, plays an important role in angiogenesis. To study the mechanisms underlying the effect of miR-21a-3p in TCs on angiogenesis induction, PI3K subunit expression was first examined. The mRNA levels of the Class I PI3K isoforms PIK3CA, PIK3CB, and PIK3CD did not significantly change with LPS stimulation. However, the protein level of p110α in TCs was significantly increased with LPS stimulation and decreased with miR-21a-3p inhibitor co-treatment. The PI3K signalling molecules AKT, mTOR, and PTEN were unaffected by either LPS or miR-21a-3p. These results indicated that PI3K signalling might participate in angiogenesis via the p110α isoform Fig. .Fig. 6ExThe proliferation of EOMA cells was then estimated after co-culture with TCs pre-treated with the miR-21a-3p or PI3K p110α inhibitor. Culture medium from TCs stimulated with LPS promoted EOMA cells proliferation, as determined by the CCK8 assay. Compared with medium from NC TCs, culture medium from TCs with miR-21a-3p inhibition significantly reduced EOMA cells proliferation Fig. a. The efThis study reports that TCs culture medium can alleviate ARDS in mice probably via angiogenesis-associated factors regulated by miR-21a-3p. TCs exposed to LPS exhibited increased miR-21a-3p expression and VEGF production, which further promoted vascular endothelial cell proliferation. The protective effects of TCs mediated by miR-21a-3p might be regulated through PI3K (p110α)/AKT/mTOR signalling and the expression levels of the downstream targets E2F8 and Notch 2 /AKT/mTOR signalling and further demonstrates the key roles of VEGF in TCs. The E2F8/Notch2 signalling might also participates in this process. These findings shed light on miR-21a-3p in TCs as a new therapeutic target for vessel protection.Additional file 1: Figure S1. The morphology of TCs. The pictures were gathered by Cell-IQ every 8 h. The white arrow showed the typical telopode.


To report on a retrospective cohort study on the effects of expanding inclusion criteria for application of cochlear implants (CIs) on the performance 1-year post-implantation.Based on pre-implantation audiometric thresholds and aided speech recognition scores, the data of 164 CI recipients were divided into a group of patients that fulfilled conservative criteria , and the remaining group of patients that felt outside this conservative criterion. Speech recognition scores (in quiet) and quality of life (using the NCIQ) of both groups, measured at 1-year post-implantation, were compared.The group that felt outside the conservative criterion showed a higher phoneme score at 1-year post-implantation compared to the conservative group, suggesting that relaxed criteria have a positive influence on the speech recognition results with CI. With respect to quality of life, both groups significantly improved 1-year post-implantation. The conservative group showed a higher benefit on the advanced perception domain of the NCIQ. Based on their worse pre-implantation hearing, this was expected.The data suggest that relaxation of CI indication positively affects the speech recognition performance of patients with severe hearing loss. Both groups of patients showed a positive effect of CI on the quality of life. This benefit relates to communication skills and the subjective day-to-day functioning in society. Cochlear implantation is a treatment for patients with deafness who do not profit from conventional hearing aids. A cochlear implant (CI) transforms the acoustic signal into an electrical signal that activates directly the auditory nerve fibers. In adults with acquired deafness, CIs provide open-set speech understanding in the majority of patients .The audiological inclusion criteria for cochlear implantation differ over countries –5. SinceThe shift of  inclusion criteria positively affects the overall performance with a CI , which mSo, what is the best CI indication and how strict should this be applied? There is a wide variability in CI indication criteria across the countries , 14–18. This means that if there is insufficient benefit of acoustic hearing aids, a patient might become a candidate for CI, even when the hearing thresholds are not at a profound level. This is in contrast with some other countries that hold on to a more conservative approach. In Belgium, for example, the inclusion criterion is set at a phoneme scores of 30%  or less and a hearing threshold  above 85 dB HL .In several countries, audiometric hearing thresholds are the sole basis for CI inclusion; however, these thresholds do not always reflect the actual problems faced by an individual with severe hearing impairment , 19. TheAnother pitfall in CI indication is that criteria are often applied rigidly. Hearing loss might be progressive (e.g. genetic types) and, therefore, it is often not the question if the patient will receive an implant but rather when the patient will be implanted. Strict inclusion based on hearing thresholds might result in postponing cochlear implantation leading to a non-optimal result owing to auditory deprivation, while the patients’ level of social functioning remains limited until better hearing is achieved using CIs.To study the effect of expanding the inclusion criteria, a retrospective cohort study is performed to analyze the effect of pre-implantation hearing level on CI performance.Based on pre-implantation audiometric thresholds and speech recognition scores obtained with well-fitted conventional hearing aids, the data of a large group of CI recipients were divided into two groups: those patients that fulfilled conservative criteria  and the remaining group of patients that felt outside the conservative criterion but still inside the broadened Dutch inclusion criterion.For the comparisons, the speech recognition scores (assessing the primary outcome of cochlear implantation) and quality of life were studied, as obtained before the intervention  and 12 months post-implantation.The pre-implantation and 12 months post-implantation results of adults with acquired severe/profound hearing loss, unilaterally implanted, were analyzed. All included patients in the database consecutively received a Nucleus CI at the Radboudumc, Nijmegen (The Netherlands), between 2010 and 2016. To be considered for cochlear implantation, the hearing loss had to be severe to profound and the obtained speech recognition measured with a well-adjusted  conventional BTE hearing aid-had to be less than 70% phoneme score .The inclusion criteria for this retrospective analysis were age at implantation > 17 years and postlingual onset of hearing loss. The exclusion criteria were abnormal anatomy of the cochleovestibular system and known psychiatric diseases. This resulted in a study group of 164 adult CI recipients . The audiological data consisted of the unaided audiometric thresholds of both ears and aided speech recognition scores, obtained with Dutch lists of monosyllables , presented in the sound-fields at 65 dB SPL .Quality of life data were measured with the standardized and validated Nijmegen Cochlear Implant Questionnaire, NCIQ , 26. TheIn Table Based on the pre-implantation audiometric thresholds and speech recognition scores of the ear to be implanted, CI recipients were divided into two groups: those that fulfilled the conservative criteria at implantation  > 85 dB HL and phoneme scores with hearing aids < 30%), the inside conservative criterion group (IC), and the outside conservative criterion group (OC). The latter group comprised all CI users that fulfilled the broadened criteria but not the conservative criteria.The two research questions that were studied wereIs the 1-year post-implantation speech recognition score of the expanded criteria group (OC group) comparable with the 1-year post-implantation speech recognition score of the conservative criteria (IC) group?Is the 1-year post-implantation quality of life score of the OC group comparable with the 1-year post-implantation quality of life score of the IC group?© SPSS© Statistics for Windows, version 22) tested using the Student T test or the Welch . Average scores are presented as mean (± standard deviation). In addition, if there was no normal distribution of the data, the bootstrap method was applied.The differences between the OC and IC subgroups were, if applicable, statistically . This difference was significant , suggesting that the relaxed criteria have a positive influence on the speech recognition results post-implantation. Table As the bootstrap method showed comparable results, the results of the Question 2. Is the 1-year post-implantation quality of life score of the IC group comparable with that of the OC group?p < 0.05) improvement after 1-year CI use was found.The average changes (and standard deviation) of the NCIQ domain scores are given in Table t = 2.07; df = 43; p = 0.045; 95% CI 0.25; 19.66). This is expected since patients that fall within IC group have a more severe hearing loss and a worse speech understanding pre-implantation and thus had more “room for improvement” with CI than patients with better hearing pre-implantation.Comparing the difference score (pre–post) between the inside and outside group, there is a trend in which the inside criterion groups improve a little more than the outside group; however, this difference is only significant in the advanced perception domain , indicating a consistency between the measured data and the patient experiences.In Fig. The results described above endorse the idea that in countries that hold on to more conservative criteria for CI candidacy, expansion of CI indication is beneficial for long-term benefit , 28, 29.Concerning quality of life, it was expected that patients falling within the conservative inclusion criteria would experience a higher benefit from cochlear implantation than the patients with some functional residual hearing (outside criterion group). This was indeed the case concerning the advanced speech perception domain; however, not with regard to speech production and the patients’ psychosocial functioning. In these domains, the benefit was found to be comparable.The speech recognition results are in line with the literature, suggesting that waiting too long before CI might increase the risk of auditory deprivation. It remains debatable what the exact criteria should be to justify both residual hearing on the one side and need for improved hearing on the other side. Based on the literature , 8, 16 aAs the present study focuses on the indication criteria for bilateral severe to profound hearing loss, the results do not reflect indication criteria for single-sided deafness (SSD) or asymmetric hearing loss (AHL). However, even though SSD and AHL are not the scope of this study, the known effect of non (or too late)-treatment of SSD or AHL  should bA limitation of this study is the retrospective study design, causing a risk of bias and confounding. A second limitation is the fact that we did not systematically test the effect of bimodal fitting, but only included the best aided condition post-implantation, based on our focus on the actual hearing situation of CI recipients.Furthermore, it should be noted that in some countries, the CI indication has recently been reconsidered (e.g. the UK) or is not as strict as the criterion for non-aided PTA and aided speech recognition applied in the present study. Nevertheless, the present retrospective data substantiate the importance of timely intervention in adults with some residual hearing to improve their communication skills. This result might stimulate a critical evaluation in case of conservative CI indication.Summarized, the data above suggest that the expansion of indications has a long-term positive effect on the speech recognition performance of patients with severe hearing loss. It affects quality of life positively (owing to an earlier change from BTE to CI). In several countries, such patients are currently not considered for cochlear implantation. The benefit relates to communication skills and the subjective day-to-day functioning in society.


Musical hallucination carries no diagnostic significance on its own. However, it is an interesting phenomenon which can occur in various organic and psychiatric disorders. We report three patients who presented with musical hallucinations due to different aetiologies, namely, due to hearing impairment, intracerebral haemorrhage, and schizophrenia. The case series also highlights the fact that different aetiologies should be managed differently for the patients to be benefited. Musical hallucinations are rare but fascinating phenomena in psychiatry which have not been explored adequately. A musical hallucination is a type of auditory hallucination where music is perceived without an external source. It is observed in primary psychotic illness, in sensory deprivation states like hearing impairment and organic psychosis.A 78-year-old widow presented with hearing of songs for a duration of six months. She identified them as vocals with the background music of a popular singer whom she adored as a young girl. Initially it was not disturbing and she enjoyed it. However, the music became very disturbing as it got louder with time. She reported gradual hearing impairment for 2 years. In neurological examination bilateral hearing impairment was detected which was confirmed to be sensorineural auditory impairment on audiography. Her mental state examination was normal with no other psychotic phenomena or cognitive impairment.The consultant otolaryngologist recommended her hearing aids after which the hallucinations disappeared completely without any psychotropics. One year following the intervention, she remains asymptomatic.A 29-year-old female was referred by the neurosurgical team as she complained of hearing music for one-week duration. She was admitted with the complaints of severe headache of sudden onset and was found to have intraventricular and intracranial haemorrhages. Following the surgical evacuation of the haemorrhages she made a full recovery. Hearing of music started one week following the surgery. She was hearing familiar songs in increased volume with distorted sounds. She had no hallucinations of any other modalities and nor there were any delusions. Her consciousness was clear and she was oriented in time, place, and person. There were impairments in her short-term memory and long-term memory along with frontal lobar impairment. She had no neurological deficits in the physical examination. A Noncontrast CT (NCCT) brain following the onset of musical hallucinations revealed no abnormalities. She was started on quetiapine 25 mg and was gradually titrated up to 150mg for which she responded. One year following the initiation of treatments she remains symptom free.An 86-year-old lady complained of being persecuted by her neighbours for 3-month duration. She reported that 2 males are living inside her body controlling her activities. She heard persistent voices chanting “pirith” (a form of religious sermon) for a duration of 2 years. The sounds were slow and rhythmic in nature. She found great relief with the voices and believed that she has achieved a higher spiritual status. In addition, she had persecutory delusions, delusions of control, and somatic hallucinations. Her cognitive functions were normal. She was diagnosed with very late onset schizophrenia and was commenced on risperidone 2mg in the night which was gradually increase to 5mg in the night. Even though her symptoms gradually improved, she regretted it as she no longer heard “pirith” chanting. She had no hearing impairment. One year following the initiation of treatments she has no symptoms.Musical hallucinations are a form of auditory hallucinations characterised by hearing of songs, tunes, and melodies in the absence of auditory stimuli . This phAs opposed to other forms of auditory hallucinations, musical hallucinations are perceived as a pleasant experience by most patients . This waIn most reported cases, musical hallucinations were found to have some personal significance to the patients . In a prThis personal significance and enjoyable nature of the hallucinations have cast doubt about the true nature of the musical hallucinations. Hence, Keshavan et al., in 1992, argued that musical hallucinations are due to memories that patient find difficult to unlearn . HoweverMusical hallucinations are known to have heterogeneous aetiologies. Hearing impairment, psychosis, organic conditions including epilepsy, brain tumours, head injury, encephalitis, multiple sclerosis, and substance intoxication are among the commonest causes. Golden et al., in 2015, identified causes of musical hallucinations in a sample of 393 patients identified by the Mayo Medical Record Linkage System . In thisThe number of cases with auditory Charles Bonnet Syndrome with hearing impairment has been reported. Lesions anywhere in the auditory tract leading to hearing impairment can lead to musical hallucinations . ReleaseStructural brain lesions can cause musical hallucinations . It has Musical hallucinations are a rare form of psychopathology in primary psychotic disorders. In a survey by Golden et al. in 2015 the majority of the patients with psychiatric causality for musical hallucinations had depression followed by bipolar affective disorder. Schizophrenia was found in 2% of patients . ContrarThere is a paucity of evidence on treatment of musical hallucinations. Treating the aetiology has been known to remit musical hallucinations in some cases. Auditory Charles Bonnet Syndrome is known to remit following the improvement of hearing . Case stThere is a deficiency of literature on this fascinating phenomenon. Further research is needed to look for the aetiology, neurobiology, and treatment of this rare but unique psychopathology.


Estimating the incidence of antiretroviral discontinuations due to adverse drug reactions (ADRs) is important to inform antiretroviral treatment (ART) regimen recommendations, and to guide prescribing and monitoring policies. Routinely collected clinical data is a useful source of pharmacovigilance data. We estimated the incidences of first-line antiretroviral discontinuations due to ADRs using routine clinical data, and compared them with incidences estimated using data enhanced by folder review, in two South African cohorts.We included patients 16 years and older on first-line ART. We selected a stratified random sample of 25% for checking of ART prescription data and reasons for antiretroviral discontinuations retrospectively, including folders reviews where required (enhanced-data sample). We estimated the incidence of antiretroviral discontinuations using Kaplan-Meier and competing risk analyses.In 15396 patients, 40% had a first-line antiretroviral discontinuation by three years on ART. We could determine the reason for 65% of discontinuations using routine data only, and 84% of discontinuations, in the enhanced-data sample of 3837 patients. ADR was the most common reason for discontinuations. In the enhanced-data sample, the cumulative incidence of discontinuations due to ADRs by three years was 30.4% (95% CI: 24.4–36.6) for stavudine; 2.0% (95% CI: 1.5–2.6) for tenofovir, and 1.3% (95% CI: 0.8–2.1) for efavirenz. Using routine data only, the cumulative incidences of discontinuations due to ADRs by three years for stavudine, tenofovir, and efavirenz respectively were 23.9% (95% CI: 20.3–27.7), 1.2% (95% CI: 0.9–1.4) and 0.5% (95% CI: 0.3–0.7).Although the relative rankings were similar using routine or enhanced data, lack of checking for missing reasons for discontinuation resulted in underestimates of the incidence of antiretroviral discontinuations due to ADRs. Systems to improve data collection of reasons for regimen changes prospectively would increase the capacity of routine data to answer pharmacovigilance questions. Adverse drug reactions (ADRs) caused by antiretroviral drugs can cause substantial morbidity and can result in compromised adherence to treatment. In orderSeveral previous African studies have used routine cohort data to describe antiretroviral discontinuations due to ADRs.–9 In allWe estimated the drug-specific incidence of first-line antiretroviral discontinuations due to ADRs using routine clinical data from two large South African cohorts. We compared the incidences with those estimated in a sample of patients where we checked reasons for antiretroviral discontinuations retrospectively. We also identified risk factors for antiretroviral discontinuations due to ADRs.We included all antiretroviral-naïve patients who were at least 16 years old and started antiretroviral treatment from 01 January 2010 until 31 December 2012 (Themba Lethu) or 15 October 2013 (Khayelitsha). Khayelitsha HIV Treatment Programme, which comprises three primary healthcare clinics, and Themba Lethu Clinic, which is based at a secondary hospital, both manage patients according to standard ART guidelines published by the National Department of Health, which follow the World Health Organization recommendations. Both sites prospectively collect routine clinical data, including demographic details, antiretroviral prescriptions, and laboratory results, electronically.The CD4 count threshold for ART eligibility changed from <200 cells/μL to <350 cells/μL in April 2013. RecommenAnonymised data were exported from both sites’ electronic databases using a standard data transfer format. We checked for data inconsistencies and possible errors, which were sent to the sites for resolution. In addition, we selected a stratified random sample  of 25% of patients overall for detailed checking of ART prescription data against clinic records, and to identify reasons for antiretroviral discontinuations where those were missing, through retrospective folder reviews where required (‘the enhanced-data sample’). The routine-data group comprised all patients not included in the enhanced-data sample.We defined antiretroviral discontinuations as discontinuation of at least one first-line antiretroviral drug for any reason, regardless of whether or not another drug was substituted. Reasons for discontinuations were determined at the time of discontinuation by the treating clinician, and included adverse drug reactions, treatment failure, contraindication (including pregnancy and drug interactions), or guideline change. Causality assessment of adverse drug reactions was based on clinical judgement of the treating clinician. In addition, discontinuations of at least two drugs (including a switch from a non-nucleoside reverse transcriptase inhibitor to a protease inhibitor), after at least one viral load >1000 copies/mL, that occurred at least six months after treatment initiation were also classified as treatment failures, provided that no other reason was given by the treating clinician.We estimated the incidence of antiretroviral discontinuations for the five most-commonly prescribed antiretrovirals using Kaplan-Meier analyses. We estimated the drug-specific incidence of discontinuations for different reasons using competing risks analysis. We explored associations between discontinuations due to adverse drug reactions and treatment site, sex, age, and baseline weight, CD4 count, and estimated glomerular filtration rate using competing risks regression. Antiretroviral discontinuations for reasons other than toxicity were considered competing risks. Patients were censored at death, loss to follow up, transfer to another treatment facility, or analysis closure (six months before database closure at each site). Patients were considered lost to follow up if they had no clinic visit after analysis closure (six months before database closure).The University of Cape Town and the University of the Witwatersrand Human Research Ethics Committees approved the sites’ contribution of anonymised data to the study. The University of Cape Town Human Research Ethics Committee approved the analysis. The study protocol was also reviewed and cleared as human subjects' research by the U.S. Centers for Disease Control and Prevention (CDC).We included 15 396 patients: 9 630 from Khayelitsha, and 5 766 from Themba Lethu. Their characteristics at the time of ART initiation are summarized in Overall 1 832/15 396 patients (12%) had an antiretroviral discontinuation. The incidence of antiretroviral discontinuation for any reason was 110.3 per 1 000 patient years  105.3 to 115.4). Of those remaining in care for at least three years, 298/495 (60%) remained on their initial regimen at three years. This differed according to ART regimen: 81/99 (82%) of those on any tenofovir-based regimen (predominantly combined with efavirenz and either lamiviudine or emtricitabine), and 128/262 (49%) of those on any stavudine-based regimen were still on their original regimen at three years .The reason for antiretroviral drug discontinuation was unknown in 35% of patients in the routine-data group, and 16% in the enhanced-data sample . The incidence of antiretroviral discontinuations due to ADRs was 28.2 (95% CI 25.4 to 31.3) per 1 000 patient years in the routine-data group, and 45.9 (95% CI: 39.8 to 52.9) per 1 000 patient years in those patients in the enhanced-data sample.In the enhanced-data sample ADR was the most common reason for antiretroviral discontinuation, followed by treatment failure: 12.1% and 10.0% of patients had an antiretroviral discontinuation because of toxicity or treatment failure respectively by 3 years on ART. The proportion of antiretroviral discontinuations  attributable to the various causes is shown in The cumulative incidence of antiretroviral discontinuations due to ADRs according to each antiretroviral is shown in The majority of tenofovir discontinuations due to ADRs were for kidney toxicity . In a further 11 patients (20%) kidney toxicity was the likely reason, as they had a decrease in estimated glomerular filtration rate (eGFR) (in 8 patients to less than 60 mL/min) before tenofovir was discontinued.Stavudine discontinuations due to ADRs were for lipodystrophy , peripheral neuropathy , and hyperlactataemia ; the toxicity was unspecified in 11 patients. The majority of zidovudine discontinuations due to ADRs were for haematological toxicity (9/13 patients).Efavirenz discontinuations due to ADRs were for hypersensitivity , neurological/neuropsychiatric toxicity , and hepatotoxicity in 1 patient. Nevirapine discontinuations due to ADRs were for hypersensitivity (9/16), or hepatotoxicity (7 patients).In a multivariable analysis of the enhanced-data sample patients, tenofovir, stavudine, and nevirapine discontinuations due to ADRs were associated with treatment site . In addiIn our cohort of 15 396 patients, 40% had a first-line antiretroviral discontinuation by three years on ART. In the enhanced-data sample of 3 837 patients, we were able to determine the reason for 84% of discontinuations. ADR was the most common reason for antiretroviral discontinuations, followed by treatment failure, with 12% and 10% of patients having discontinued antiretrovirals for those reasons respectively by three years on ART. The incidence of discontinuations differed by antiretroviral drug: 30% and 2% of patients had stopped stavudine or tenofovir respectively because of ADRs by three years on ART. Although the relative rankings of antiretroviral discontinuations were similar using routine data only, lack of additional checking for reasons resulted in underestimates of the incidence of antiretroviral discontinuations due to ADRs. Standard first line antiretrovirals tenofovir and efavirenz had the lowest incidences of discontinuations due to ADRs. The majority of tenofovir discontinuations were due to decreases in kidney function, and the majority of efavirenz discontinuations were due to hypersensitivity reactions or neuropsychiatric symptoms. Associations with tenofovir discontinuations due to ADRs included older age, and lower baseline CD4 count, weight, and estimated glomerular filtration rate. We found no associations between efavirenz discontinuations due to ADRs and potential risk factors that we considered. In general, discontinuations for ADRs were more likely to occur at the hospital-based site (Themba Lethu) than the primary health care clinic (Khayelitsha).We found an incidence of discontinuations due to ADRs of 45.9 (95% CI 39.8 to 52.9) per 1 000 patient years. This is slightly lower than estimates from other African cohort studies that were conducted before the 2009 World Health Organization guideline change that replaced stavudine with tenofovir as part of the preferred first-line ART regimen. A study in Cote D’Ivoire reported an incidence of toxicity-related treatment modifications or interruptions of 5.2  per 100 patient years. A study Using only routine data identified relative trends in antiretroviral discontinuation rates and identified relevant risk factors for antiretroviral discontinuations due to ADRs; but substantially underestimated the absolute incidence of discontinuations due to ADRs. The retrospective data checking process was labour intensive, and involved a substantial amount of time, both for generating queries, and for site staff to perform folder reviews, but reasons that were not recorded in the database were frequently available in the folders or other patient records. This suggests that improving prospective data capture systems in routine databases would increase the capacity of routine data to answer pharmacovigilance questions.Patients at Themba Lethu, the hospital-based cohort, were more likely to have tenofovir and stavudine discontinuations due to ADRs than patients at the Khayelitsha cohort primary health care clinics. Patients at the hospital site had more frequent laboratory monitoring, and healthcare workers there were more likely to have access to blood results sooner, which might in part explain the difference. These differences in the patterns of reasons for antiretroviral discontinuations highlights the importance of using data from several representative sites in order to make conclusions regarding ADR-related antiretroviral discontinuations.Our study had several limitations. Data were from routine clinical practice, and even after thorough data checks there were still missing data, including unknown reasons for antiretroviral discontinuations. Reasons for discontinuations were as recorded by the treating clinician. We might have underestimated the rates of ADR-related discontinuations if the reason wasn’t adequately recorded. We assessed only the first antiretroviral discontinuation, so might have underestimated rates of discontinuations that occurred later on in the course of treatment. Similarly, we might have underestimated antiretroviral discontinuations owing to patients being lost to follow up.Currently recommended first-line antiretrovirals were relatively well-tolerated in our setting.The routine data used in our study was adequate to estimate relative trends in, and identify potential risk factors for, antiretroviral discontinuations due to ADRs. But obtaining more accurate incidence estimates involved a time- and labour-intensive data-checking process. Observational cohorts are a useful source of pharmacovigilance data. Improving prospective data capture systems would increase the capacity of routine clinical data to answer pharmacovigilance questions.S1 Table(PDF)Click here for additional data file.


Batch experiments were completed to assess the sludge reduction of the metabolic uncoupler 3,3′,4′,5-tetrachlorosalicylanilide (TCS). The effects of various TCS concentrations on sludge yield were evaluated and the mechanisms associated with sludge reduction were assessed. We discovered that TCS addition resulted in a reduction in sludge. Furthermore, a low dose of TCS (≤3 mg/L) resulted in a slight reduction in the efficiency of the wastewater treatment system, while >3 mg/L TCS reduced matrix removal efficiency, with an especially remarkable inhibition effect on ammonia removal. An increase in TCS addition was associated with a gradual decrease in both the electron transport system (ETS) activity and the specific cellular ATP (SATP) in the TCS system. It was demonstrated that TCS plays an important role in metabolic uncoupling. However, with the addition of TCS, both contents and compositions were increased, and the protein content increased more than polysaccharide production in extracellular polymeric substances (EPS). At TCS concentrations of ≤3 mg/L, DNA content was stable, but it increased rapidly from 4.97 mg/L to 15.34 mg/L as the TCS concentration was elevated from 6 mg/L to 12 mg/L. This implied that the mechanisms of sludge reduction were different for different TCS concentrations, including uncoupling metabolism, maintenance metabolism and lysis–cryptic growth. Conventional activated sludge is a broadly utilized process in both municipal and industrial wastewater treatment plants (WWTPs). One disadvantage of this process relates to the significant quantities of excess sludge that are generated, which in some cases are as high as 50–60% of the total operational expenses of the wastewater treatment plants . Post-trOf the above-mentioned in situ excess sludge reduction technologies, the metabolic uncoupler is most widely accepted, since it can seamlessly feed into the aeration tank of a wastewater treatment plant, which avoids the necessity to change the conventional wastewater treatment process; because the metabolic uncoupler is usually a lipophilic soluble acid, it can transport protons across the cell membrane and result in partial decomposition due to catabolism and anabolism ,13,14,15The majority of studies on metabolic uncouplers have been aimed at determining their capacity to decrease biomass production and the impacts of chemical uncouplers on substrate removal efficiency ,13,14,15In the current evaluation, the effects of TCS addition on extra sludge yield and nutrient elimination were evaluated. Furthermore, the impact of TCS on microbial activity inhibition and energy distribution was also investigated. We believe that the findings will help improve our comprehension of the reaction of activated sludge systems to metabolic deodorants, such as TCS in biological wastewater treatment.The activated sludge utilized in the experiments was sampled at a local municipal wastewater treatment plant, following which it was sieved (2-mm mesh) to eliminate coarse pieces. Prior to experimentation, the sludge was cultivated at room temperature in six identical sequencing batch reactors (SBR), each with a working volume of 3.2 L. The SBRs were operated for four cycles daily, and every cycle lasted 6 h. There were five stages in each cycle: fill (30 min), react (4 h with aeration), settle (1 h), draw (20 min), and idle (10 min). For the filling, drawing, and aeration reactions, the operating cycle was controlled automatically with a time device. The dissolved oxygen (DO) concentration was maintained greater than 4 mg/L, and the mixed liquid suspended solid (MLSS) level was retained at around 2500 mg/L by removing any extra sludge. The sludge was cultivated for 40 days in the absence of TCS, following which different concentrations  of TCS were added.The synthetic wastewater utilized for cultivation comprised sodium acetate and sucrose as the only carbon source for bacterial growth; ammonium chloride was used as the nitrogen source as well as additional trace elements, as shown in −1 h−1), D485 is the absorbance at 485 nm, V is the extract volume (mL), k is the slope of the standard curve (mL μg−1), W is the dry weight of the activated sludge (mg), and t is the incubation time (h).The homogenous sludge samples were obtained every three days to calculate the electron transport system (ETS) activity. 2,3,5-triphenyltetrazoliun chloride (TTC) is a kind of oxidoreduction dye used to detect the activity of sludge ETS. TTC-ETS was calculated to evaluate the impacts of TCS on ETS activities, and MLSS was utilized as an indicator of the biomass of sludge microorganisms. TTC-ETS activity was calculated based on Kenner Ahmed , as modiCellular ATP was removed using the altered procedure of Jiang and Liu . BrieflyA cation exchange resin (CER) strategy modified from Frolund et al.  was usedobs), the excess biomass increase was divided by the corresponding decrease in COD concentration after the addition of TCS. Yobs refers to the new amount of sludge generated for each removal of 1 g COD, that is, Yobs = ΔMLSS/ΔCOD. Sludge reduction rate (SR) were determined utilizing Equation (2):To calculate the observed biomass yield  in the control reactor without TCS, and R is the TTC-ETS activity (or SATP) in the TCS reactor.The inhibitory actions of TCS on SATP and TTC-ETS activity were assessed based on the corresponding inhibitory rates, which were determined utilizing Equation (3):obs at various TCS concentrations are indicated in obs with TCS concentrations within 0–3 mg/L. The depletion effect on the sludge was noteworthy and reached 40.13% at 3 mg/L, whereas an additional elevation in TCS content did not result in a more significant depletion in Yobs. Nearly identical findings have been documented previously [The reduction percentages of Yeviously ,14,28.4+-N removal efficiencies under various concentrations of TCS are presented in 4+-N removal efficiency show a significant deterioration in the mean removal efficiency for NH4+-N with the addition of TCS, indicating the significant inhibition of TCS on the nitrification performance. When TCS was 1 mg/L, the average NH4+-N removal efficiency decreased to 9.1%. However, a high dose of TCS in activated sludge had a remarkable influence on the NH4+-N removal efficiency, decreasing from 63.1% to 51.4% with an increase in TCS concentration from 6 to 12 mg/L. The findings corroborate those of Zhang et al. [4+-N oxidation was greatly obstructed by the addition of uncouplers. This inhibitory impact was the result of the suppression of ammonia-oxidizing bacteria (AOB) activity by metabolic uncouplers, as proven by the decrease in ammonia uptake rate (AUR) with metabolic uncoupler addition [The COD, TP, and NHg et al. , who alsaddition . The finaddition ,31.At TCS concentrations 0–3 mg/L, slightly lower COD removal efficiencies were detected than in the absence of TCS. A gradual decrease in COD removal efficiencies was observed with increased TCS concentration. TCS concentration 0-6 mg/L had little effect on TP removal, which may have a small effect on polyphosphate accumulating bacteria. TCS concentrations of 6–12 mg/L were associated with COD removal efficiencies of 81%, and they sharply decreased by approximately 8% with the addition of 3 mg/L TCS.In the literature, three mechanisms of sludge reduction, namely maintenance metabolism, uncoupled metabolism and lysis–cryptic growth, are discussed. Alterations in ETS activity, SATP, and EPS content in the activated sludge culture as well as the protein and DNA contents in activated sludge supernatants have been widely used in the evaluation of metabolic uncoupling, metabolism, and lysis–cryptic growth ,33. We m−1 h−1) was found, only decreasing by 2.54% compare to the control (131.23 µg mg−1 h−1). With a dose of 1 mg−1, the activity sharply decreased to 105.79 µg mg−1 h−1 (19.38%). At 12 mg/L TCS, the TTC-ETS activity of the microorganisms was 89.98 µg mg−1 h−1, which indicated a reduction in the sludge of around 31.43% compared to the control.ETS activity has been suggested as a particular indicator of aerobic microorganism respiration and as a suitable means of estimating the respiratory possibility of these populations . TypicalUncoupler TCS might promote the cell membrane permeability increase of H+, leading to the decomposition of phosphate coupling oxides, and, subsequently, the synthesis of adenosine triphosphate (ATP) is reduced, thus achieving sludge reduction ,13. FiguThe EPS content and composition in the existence of TCS at various concentrations are indicated in The DNA levels in bulk solution are indicative of the degree of cell lysis (Aquino and Stuckey 2004). As indicated in (1) The degree of sludge reduction was dependent on the ratio of the first uncoupler concentration to the first biomass concentration, which was MLSS:TCS of 2500:3 mg/L.4+-N by TCS.(2) TCS at low concentrations had little effect on TP removal, followed by COD, but it did have a larger effect on the elimination of NH(3) The presence of TCS can weaken TTC-ETS activity and reduce SATP synthesis, leading to inhibition of microbial activity.(4) A difference in concentration was the demonstrated mechanism of sludge reduction. High concentration (6 and 12 mg/L) involved lysis–cryptic growth in sludge reduction by TCS. At a slight concentration (≤3 mg/L), metabolic uncoupling and alterations in energy distribution were pivotal for achieving effective sludge reduction by TCS.


This report serves as a guide for investigating mucormycosis infections in healthcare. We describe lessons learned from previous outbreaks and offer methods and tools that can aid in these investigations. We also offer suggestions for conducting environmental assessments, implementing infection control measures, and initiating surveillance to ensure that interventions were effective. While not all investigations of mucormycosis infections will identify a single source, all can potentially lead to improvements in infection control. Mucormycosis is a rare but serious infection that can affect immunocompromised patients in healthcare settings. Investigations of possible mucormycosis outbreaks in healthcare settings have enabled critical insights into the exposures and environmental conditions that can lead to transmission. The purpose of this report is to compile resources and experiences that can serve as a guide for facilities and health departments investigating mucormycosis infections and suspected outbreaks in healthcare settings. We first provide an overview of mucormycosis outbreak investigations and then suggest how the typical steps of an outbreak investigation can be adapted and applied to mucormycosis infections in hospitals.Rhizopus and Mucor species, but others include Apophysomyces, Rhizomucor, Cunninghamella, Lichtheimia, Cokeromyces, and Saksenaea [Mucormycosis is caused by a group of molds called mucormycetes, which are found in the environment and release spores that are easily aerosolized and dispersed . The mosaksenaea . Mucormyaksenaea .Because mucormycetes grow best in acidic, high-glucose environments, diabetes, especially if poorly-controlled or complicated by ketoacidosis, is also an important risk factor . Some evExposure to mucormycetes can cause outbreaks among at-risk patients in healthcare settings, with infections linked to construction, water leaks, insufficient air filtration, non-sterile medical supplies, deficient device reprocessing, dental procedures, contaminated linen, contaminated food and supplements, and other sources of mold. These infections are often devastating, with reported mortality rates around 50% in some studies ,14,15. TBecause mucormycosis is neither nationally notifiable nor reportable in any US state, there is no public health surveillance system to monitor infections, so it is difficult to discern whether a hospital’s infection rate is above average. Mucormycosis is a rare infection even among high-risk patients, with a 12-month incidence of <1% in one cohort of hematopoietic cell transplant (HCT) and solid organ transplant recipients during 2001–2006  and an iA final challenge in investigating mucormycosis outbreaks is that mold spores can be found in many different environmental sources, including air, dust, soil, and on surfaces, and may not be visible. Infection control assessments often identify multiple areas for improvement, and it is rare to identify a single point source of mold that is epidemiologically linked to cases. Because many outbreaks are multifactorial and require hospitals to take several mitigating actions simultaneously, it can be impossible to assess the effects of individual interventions. The most widely used and accepted case definition for invasive fungal infections was developed for use in clinical trials among transplant patients by the European Organization for Research and Treatment of Cancer (EORTC) and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (MSG) . CDC hasCase definitions for cutaneous infections have included soft tissue infections with evidence of mucormycosis, such as positive fungal culture and observation of broad pauciseptate hyphae with branching at right angles on microscopy, mainly at the periphery of the skin lesion . Case deLaboratory confirmation of mucormycetes in patient specimens is a central part of all case definitions above, although mycology laboratory capacity varies by hospital. Those with mycology expertise should identify species of suspected mucormycete cultures, although species identification of mucormycetes can be difficult, and misidentifications occur commonly in clinical laboratories. Where capacity is limited, cultures may be sent out to reference diagnostic laboratories. In some states, public health laboratories can assist with diagnostics. The CDC offers a comprehensive fungal diagnostic service and can typically receive suspected mucormycete isolates when state public health partners are involved . PolymerBecause mucormycosis is rare even among patients at the highest risk, investigation is warranted even when it is unclear whether an increase in the number of infections represents a true outbreak, particularly if cases appear to cluster in time or place or among patients who share other exposures. For example, an increase from two to four cases per year at a single hospital could be because of chance or could signal that a new source of mucormycetes is present in the facility, leading to infections. Aspergillus, which causes infections more frequently than mucormycetes. Tracking Aspergillus infections may help to identify problematic exposures, because like mucormycosis, these molds are transmitted via airborne spores. Facilities may consider looking for infections in the previous 12 to 24 months by reviewing microbiology records for positive fungal culture results, and, if possible, reviewing histopathology specimens with evidence of tissue invasion by fungal hyphae. A spreadsheet of specimens  should be created that includes basic information about each culture or pathology specimen, including a patient identifier, date collected, and organism identified. This initial line list can be expanded to include additional clinical details as the investigation progresses , as well as discharge dates for previous hospitalizations within several months of suspected onset;Prior admissions and outpatient visits;Patient risk factors and underlying conditions ;Immunosuppressive medications ;Signs and symptoms of fungal infection, with approximate dates of onset;Histopathology specimen collection date(s) and findings;Culture date(s), specimen source, and species;Antifungal medications given with dates and indication (prophylaxis versus treatment);Unit and room assignments with dates;Invasive disease versus colonization by EORTC/MSG definition or clinician determination;Devices, procedures, lines, and surgeries;Outcomes, including date of discharge or death.Key details that should be collected on a line list for each patient include: Visualizing the location of patients in relation to events such as construction, renovations, maintenance work, and water and air leaks may help to identify shared exposure. Maps of the air handling systems with relative pressures between units, hallways, and rooms can be overlaid with patient information to determine whether locations shared by cases were supplied by the same air handlers. The frequent movement of patients within a healthcare facility can complicate mucormycosis investigations. In investigations with complex patient histories, it can be helpful to create a timeline of each patient’s location, including room and unit, in the months prior to diagnosis . This maConsidering whether transmission more likely occurred in the healthcare setting or in the community is important. Because the incubation period for mucormycosis is not well established, no standard definition of healthcare-associated mucormycosis exists. However, some evidence is available about the incubation period of cutaneous infections. A review of cutaneous mucormycosis infections found that the median incubation period for 168 patients with a single percutaneous exposure was 8 days (interquartile range: 5–16 days) , similarThe incubation period for rhinocerebral and pulmonary mucormycosis is not well established, making it difficult to establish a time period in which to identify relevant exposures. Patients at high risk for healthcare-associated mucormycosis may have had numerous procedures and inpatient stays in many rooms on different units. One strategy that may be used when evaluating patients with long hospitalizations and multiple healthcare exposures is to begin by focusing on exposures in the 14 days before suspected infection onset, then expanding to a longer window before infection if common exposures to possible sources of mucormycetes are not evident. Consider using a longer window for infections when the time of onset is difficult to pinpoint. For example, in transplant patients, the time since transplant can guide exposure identification.The descriptive epidemiology described above can provide clues to common exposures among patients and enable hypothesis generation. In However, outbreak investigations linking infections to a single exposure are the exception rather than the rule; most investigations lack sufficient evidence to attribute a mucormycosis cluster to one point source. CDC records include 17 mucormycosis-related consultations from January 2005–April 2019, of which six resulted in an epidemiologically robust attributable exposure ,41,42,43Rhizopus infections in at least 11 hospitals [Rhizopus species were isolated from unused or partially used bandages in multiple hospitals [Cutaneous mucormycosis infections have been associated with the use of non-sterile bandages, patches, tape, and adhesives, with the first necrotic lesions usually appearing underneath the product. Between 1977 and 1978, elastic adhesive bandages were implicated in an outbreak of more than 20 ospitals ,46,47,48ospitals ,50. The ospitals . Lichtheimia corymbifera from non-sterile bandages. Both hospitals subsequently recommended using only sterile bandages for deep, traumatic, and extensive wounds, including burns [L. corymbifera. Of those seven patients, five developed infections and three died. A third hospital in France used real-time PCR to test non-sterile bandages used in the burn unit, detecting Lichtheimia spp. in six out of eight crepe bandages and four out of six elasticized bandages [Lichtheimia spp. Two hospitals, one in Belgium and one in Pakistan, had outbreaks of ng burns ,52. Patibandages . The hosUlcers caused by mucormycosis found under tape used to secure an endotracheal tube were reported at two different hospitals, although neither tape was tested for contamination ,54. As aRhizopus arrhizus (oryzae) at the site of recent surgery to create a stoma [Rhizopus arrhizus was isolated from 10 of 18 unopened karaya ostomy bags at the hospital, and from 3 of 29 karaya ostomy bags from other hospitals. The median time to first ostomy bag change was 8.5 days for the case-patients, compared with 1.5 days for patients who underwent the same procedure but did not develop an infection; the hospital replaced karaya gum with a synthetic product and recommended more frequent changes. An earlier case report describes an infection that developed around an ileostomy, but the ostomy supplies were not cultured [Adhesives used to secure ostomy bags can also be considered as a source of infection, particularly if they are infrequently changed. In 2005, two patients at the same hospital developed skin and soft tissue infections with  a stoma . The adhcultured .Other cutaneous mucormycosis infections have been linked to adhesive patches used to attach a temperature probe  and hearRhizopus delemar, at a children’s hospital over 11 months found that clean linens and clean linen delivery bins from the off-site laundry were contaminated with the same species [The investigations described below suggest that investigators of possible mucormycosis outbreaks should consider a comprehensive evaluation of hospital linen washing, transportation, storage and use, from the laundry facility to the patient. Fungal outbreaks from linen have been attributed to contamination of laundered textiles at off-site laundries, during transportation, and during storage in hospitals . An inve species . The chiRhizopus spp. from multiple laundry carts. The outbreak stopped when the hospital changed its practice from cleaning carts only when they were soiled to a routine cleaning schedule, as recommended by the Healthcare Laundry Accreditation Council [In another investigation of skin and soft tissue infections that began in the axillae of four immunocompromised patients on the same unit, the facility recovered  Council .Rhizopus microsporus [Another hospital found widespread contamination of hospital linen after six patients in less than two months had infections with rosporus . Two infIn response to a mucormycosis outbreak where linens were one suspected source, investigators conducted an audit of 15 transplant and cancer hospitals and found seven (47%) centers had freshly laundered healthcare linens contaminated with mucormycetes. Three (20%) of the 15 centers failed to meet hygienically clean standards for mucormycetes . In totaRhizopus microsporus was detected on wooden tongue depressors used to prepare oral medications given through a nasogastric catheter [Because wood is a natural substrate for mucormycetes, any wooden products used in patient care should be investigated. Wooden tongue depressors were associated with infections in five adults diagnosed with gastric mucormycosis. catheter . Given scatheter ,97,98, tcatheter .Rhizopusoryzae following the use of a probiotic powder intended to contain three bacteria, Bifidobacterium lactis, Streptococcus thermophilus, and Lactobacillus rhamnosus. Testing showed the powder was contaminated with R. oryzae [Lactobacillus supplement [Mucor indicus isolates from the patient’s liver aspirate and the supplement. Infections have also been linked to contaminated medications, supplements, and food. In October 2014, a premature infant died of gastrointestinal mucormycosis caused by . oryzae . An earlpplement . The PCRIn response to an outbreak of seven intestinal infections over six months, a team of investigators at a hospital in Hong Kong found that the infections were associated with contaminated allopurinol tablets and pre-packaged food . While sMucormycetes can be transmitted during insertion of catheters and lines. Mucormycosis infections were reported in an adult patient with a permanent bladder catheter  and in tAnother possible exposure to consider, particularly for mucormycosis of the maxilla or sinuses, is a previous dental procedure, such as an extraction, that could allow spores to enter where the tooth was removed. In India, cases of rhino-orbital mucormycosis were reported in two patients, one diabetic, who had dental procedures 8 and 12 days before hospitalization . Case reMucormycosis infections that appear to begin near a surgical site may be the result of contamination in the operating, recovery, or patient rooms. Infections have been found six days following explantation of left ventricular assist devices (LVADs)  and up tBecause patients with poorly controlled diabetes mellitus are at higher risk for mucormycosis, any exposure that breaks the skin should be assessed as a potential source of spores. Infections have been documented in diabetic patients that began at the site of insulin infusion pump insertion , daily iMedical devices that have not been sufficiently sterilized may be contaminated with spores. One hospital ended an outbreak of postoperative bone mucormycosis among three patients after improved sterilization of screws used in ligament reconstruction .Mucormycosis among organ transplant recipients should be investigated when multiple case-patients share a common donor, when the donor or organ might have been exposed to mold, or the donor had evidence of a fungal infection. Mucormycosis infections have been reported following transplants with solid organs that may have become contaminated with mold at the time of the donor’s death, or during harvesting, shipping, or transplantation . For exaMucor sp., isolated Mucor sp. and Aspergillus sp. in dust from an air conditioner duct and in air samples. The team found that the air filter used was likely insufficient to trap spores [Insufficient air filtration and air intakes that are near the ground, outdoor construction, or exhaust, may also be associated with mucormycosis infections. A hospital investigating four cases of mycotic endocarditis among open-heart surgery patients in a single year, including one with p spores . Rhizomucor pusillus, as were the intake unit and the yard. Another air conditioning installation at the same hospital had fewer mucormycetes and no Rhizomucor pusillus; no mucormycetes were detected in air samples from two comparison hospitals in the same city. At a second hospital, a cluster of three sinus infections in leukemia patients hospitalized on the same hematology unit was also suspected to be caused by poor air filtration . The hosNegative pressure rooms, which are used to isolate patients with an airborne infection, have been identified as risk factor for mold infections when used for immunocompromised patients. In contrast to positive pressure rooms that protect immunocompromised patients by minimizing infiltration of unfiltered air, negative pressure rooms are intended to prevent the spread of tuberculosis or other airborne diseases and produce a lower pressure in the room compared with the corridor or anteroom. Depending on the airflow from the corridor or anteroom, negative pressure in these rooms may pull in air, as well as fungal spores, through small gaps or crevices in the walls or ceiling. Investigators concluded that a cluster of three probable and one suspect mucormycosis infections were likely caused by the use of a negative pressure room for immunocompromised patients . All patAspergillus infections in premature infants on a neonatal intensive care unit (NICU) adjacent to hospital construction, with inadequate barriers that allowed a higher density of mold spores measured in the unit than a comparison area without construction [Hospital construction, inside or outside the facility, may result in dispersal of dust, aerosolization of spores, and contamination of air handlers, leading to an increased incidence of fungal infections ,106,107.truction . InvestiAspergillus spores near the water-damaged wall, while the spore concentration was zero at comparison sites near the ends of the unit. The spore counts dropped to zero a week after remediation of the leak and wall with mold.Water leaks have been identified as a potential source of mold exposure. In one hospital outbreak, two children who developed mucormycosis infections on a pediatric oncology unit were in beds closest to a linen closet that shared a wall with a leaking shower . One devApophysomyces elegans was identified in a non-insulin dependent diabetic man after an allergy test that included application of a petal, stem, and leaf from a snapdragon plant to his unbroken skin under a sterile dressing [Cunninghamella species but not A. elegans; samples of the adhesive did not grow mold or fungi. Investigators hypothesized that the flower was likely contaminated by mold from the soil and recommended washing plants and using less occlusive dressings for future allergy tests, although the efficacy of these interventions is unknown. Because mucormycetes are found in soil, any patient exposure to plants, especially in a healthcare setting, should be considered among possible sources of infection during an investigation. One case of cutaneous infection with dressing . The manMucor circinelloides [Case reports have described infections acquired in the community by injection drug use , motor velloides , with onelloides .Environmental assessments are critical in developing hypotheses in response to a suspected mucormycosis outbreak. These assessments include an examination of the facility with experts in infection prevention and facility staff who oversee building, heating, ventilation, and air conditioning systems (HVAC), construction and maintenance, environmental cleaning, and linen transport and storage. Facility records can also help to identify lapses in scheduled maintenance, construction, and repair history, and other events that may have created environments that support mold growth. Investigators can check any reports to the facilities or maintenance department about possible mold, including any mold referred for remediation. Visual and records-based environmental assessments should typically be performed before conducting sampling. To guide systematic environmental assessments for investigations of mold infections in healthcare settings, the CDC developed a checklist . General inspection for mold, leaks, dirtHVAC systemsEnvironmental cleaningConstruction and maintenanceLinenThe checklist  covers tA second checklist to assess healthcare linen in more detail is published by the Healthcare Laundry Accreditation Council, a US organization that inspects and accredits laundries serving healthcare facilities .Facility work orders for routine maintenance and repairsInfection Control Risk Assessments (ICRAs) conducted before starting constructionIndoor air temperature and humidity logs, including any days when humidity exceeded 60%Dates of air filter changesHVAC system, filters, and fans maintained according to manufacturer instructionsFrequency and method of filter performanceAir flow monitoring recordsIn addition to visual inspection, review of facility records can also help investigators identify possible sources of mold. Records to review include: The CDC’s Guideline for Isolation Precautions details protective environments needed for allogeneic hematopoietic stem cell transplant (HSCT) patients . The rooOnce hypotheses have been developed from the epidemiologic and environmental review, investigators can evaluate the proportion of infected patients with each exposure before onset. If possible, it may also be useful to conduct an analysis limited to proven cases. Although some studies have used cohort and matched case-control designs to evaluate whether patient risk factors such as neutropenia are associated with invasive fungal infection incidence or mortality ,122,123,Nonetheless, epidemiologic analyses, while not necessary in all investigations, can add to the weight of evidence about exposures that increase patient risk and inform recommendations for patient care. A study of infections at the site of ostomy bags compared median times between ostomy bag changes in a cohort of patients following stoma creation and found a longer time before bag change in patients who developed infection than those who did not . A cohorHealthcare facilities are often inclined to begin environmental sampling early in an investigation. While environmental sampling may aid an investigation, it can also be expensive and misleading. Any environmental sampling should be part of a larger building assessment that includes an inspection for moisture incursion and potential sites of mold growth ,125. An Molecular epidemiologic tools such as comparing DNA in isolates from patients to isolates from the environment can be helpful in evaluating epidemiologic hypotheses. Some investigators have used whole genome sequencing (WGS) to assess relatedness of isolates , but theThus, we recommend considering environmental sampling only after examining shared exposures between cases and conducting environmental assessments including visual inspections and record review. Information obtained during case interviews can focus the environmental evaluation on potential areas of exposure. Environmental sampling methods can include the use of settle plates and spore trap samples followed by microscopic identification. These methods can often be problematic for mucormycetes as a single spore can overgrow an entire plate in less than 24 h, crowding out other species and making the extent of contamination difficult to determine. Combinations of sampling methods can be most powerful, as each method has weaknesses. For example, spore trap samples often cannot detect large, non-airborne spores, or small spores that are not pulled into the sampler due to the short sampling times. Hard surfaces can be sampled using swabs, and dust can be collected from soft surfaces like linen and carpet using small vacuums.Comparing spore counts and species profile between indoor and outdoor samples can be useful in interpreting results. The concentration of mold in outdoor samples should exceed indoor samples, and the species profiles should be similar. The presence of a predominant species in indoor air that is not found in outdoor air may indicate an indoor source that needs remediation. Because little is known about spore counts on linen or in air in the absence of an outbreak, environmental sampling to test results for the hypothesized exposure against a comparison group may be most informative. For example, studies have compared the proportion of contaminated linen or karaya ostomy bags at one hospital to proportions at other hospitals , spore cAlthough it may not be possible to pinpoint the source of mold exposure during an outbreak, improving infection control practices can prevent future healthcare-associated infections. We recommend remediating all infection prevention gaps identified in the investigation. Support from hospital administration is key to the success of these interventions. Infection prevention should ensure that a risk assessment is performed prior to any construction or maintenance activities that may disperse dust. It is also important for infection prevention to collaborate with plant operations managers for the routine inspection of the condition of barriers used during construction, as well as airflow and air filtration from the construction site. Routine rounding in patient care areas should include proactive observation for water leaks that may become reservoirs for mold. Healthcare personnel should have competency-based training in the use of both negative and positive pressure rooms, including indications for the use of these rooms. A partnership between infection prevention and environmental services managers may be needed to identify new processes for handling clean linen to avoid environmental contamination prior to patient use.Some facilities have implemented control measures that go beyond current recommendations in response to an outbreak. Some have given prophylactic antifungals effective against mucormycetes after considering the potential risks, patient tolerance, adverse effects, and cost. These extra measures have included expanding the use of respirators to patients who are immunosuppressed but did not receive HSCT, removing plants from areas outside protective environments, and using sterilized linen for some high-risk patients. To date, evidence is lacking to support or refute the efficacy of these expanded interventions for preventing mold infections in patients who have not had HSCT. Facilities considering interventions beyond the current recommendations can help expand the evidence by evaluating and publishing the results. For all interventions, prospective surveillance is needed to assess whether infections continue to occur. Ongoing, active surveillance of culture and histopathology specimens yields important information about whether interventions have been effective. Because they offer more detailed information, microbiology and pathology databases are recommended over diagnostic codes, such as International Classification of Diseases (ICD) codes. Although ICD codes may have low sensitivity, they can be used as a supplementary tool. When possible, infection control committees or outbreak review teams can include mucormycosis infections in their review. Medical record alerts may help for rapid notification. Ongoing collaborative surveillance across many hospitals in a region can also be invaluable in detecting when rates of infection are above baseline. While not all investigations of mucormycosis infections will result in discovery of a single source, all can point to important areas for improvement in hospital infection control. Applying the tools in this guide will allow hospitals and public health partners to build on experience gained by other teams that have faced these challenging outbreaks. A comprehensive review of past infections can yield evidence on how potential sources of mold exposure may have changed over time, while ongoing surveillance provides valuable evidence on which interventions are effective. Rapid communication with treatment teams and patients can save lives through early identification of mucormycosis infections. Environmental assessments with partners in maintenance, construction, and environmental services can point to potential gaps in infection prevention and identify areas for remediation. All of these actions can improve patient safety, even if the source of mold is never definitively established. Continued efforts to understand the sources of mold that cause mucormycosis infections in healthcare will help prevent future infections and protect vulnerable patients.


GJB2 & GJB6, whose mutations have played vital role in hearing impairment across many ethnicities in the world. In our present study we have focused on the role of these mutations in assortative mating hearing impaired families from south India.DFNB1, the first locus to have been associated with deafness, has two major genes GJB2 and GJB6 genes and the outcome of gene mutations were compared in the subsequent generation in begetting deaf offspring.One hundred and six assortatively mating hearing impaired (HI) families of south Indian origin comprising of two subsets: 60 deaf marrying deaf (DXD) families and 46 deaf marrying normal hearing (DXN) families were recruited for this study. In the 60 DXD families, 335 members comprising of 118 HI mates, 63 other HI members and 154 normal hearing members and in the 46 DXN families, 281 members comprising of 46 HI and their 43 normal hearing partners, 50 other HI members and 142 normal hearing family members, participated in the molecular study. One hundred and sixty five (165) healthy normal hearing volunteers were recruited as controls for this study. All the participating members were screened for variants in GJB2 gene), p.Q57R, p.E101Q, p.R104H (in GJB6 gene) were also identified in this study.The DFNB1 allele frequencies for DXD mates and their offspring were 36.98 and 38.67%, respectively and for the DXN mates and their offspring were 22.84 and 24.38%, respectively. There was a 4.6% increase in the subsequent generation in the DXD families, while a 6.75% increase in the DXN families, which demonstrates the role of assortative mating along with consanguinity in the increase of DFNB1 mutations in consecutive generations. Four novel variants, p.E42D  in assortative mating HI parental generation and their offspring.This is the first study from an Indian subcontinent reporting novel variants in the coding region of  Hearing is one of the vital sensations, which keeps humans connected with each other and the world around. Consequently, hearing loss can have a profound impact on cognitive, psychosocial and educational development of an individual. Greater part of our present day knowledge on the physiology of hearing has come from various studies on hearing loss.http://hereditaryhearingloss.org/). Despite this genetic heterogeneity across ethnicities, DFNB1 locus on chromosome 13q11–12, accounts for up to 50% of NSHL ) in assortative mating HI parental generation and their offspring. The DFNB1 allele frequencies for DXD mates and their offspring were 36.98 and 38.67%, respectively and for the DXN mates and their offspring are 22.84 and 24.38%, respectively. There was a 4.6% increase in the subsequent generation in the DXD families, while a 6.75% increase in the DXN families, which demonstrated the role of assortative mating along with consanguinity in the increase of DFNB1 mutations in consecutive generations. This study has revealed that assortative mating among the deaf may well be a cultural force, which along with consanguinity can have a profound influence on genetic variations, which in turn can lead to an evolutionary change in times to come.This is the first study from an Indian subcontinent reporting novel variants p.Q57R, p.E101Q, p.R104H in the coding region of