Brucellosis, Q fever and Rift Valley fever are considered as Neglected Zoonotic Diseases (NZDs) leading to socioeconomic losses in livestock globally, and particularly in developing countries of Africa where they are under-reported. In this study, we evaluated the seroprevalence of these 3 zoonotic diseases in domestic ruminants in Guinea from 2017 to 2019. A total of 1357 sera, sampled from 463 cattle, 408 goats and 486 sheep, were collected in 17 Guinean prefectures and analyzed by enzyme-linked immunosorbent assay (ELISA).Cattle was the species with highest seroprevalence  for the three diseases. The seroprevalence of brucellosis, mostly focused in Western Guinea, was 11.0% (51 of 463) in cattle, 0.4% (2 in 486) in sheep while no specific antibodies were found in goats. Q fever, widespread across the country, was the most frequently detected zoonosis with a mean seroprevalence of 20.5% (95 in 463), 4.4% (18 in 408) and 2.3% (11 in 486) in cattle, goats and sheep, respectively. The mean seroprevalence of RVF was 16.4% (76 in 463) in cattle, 1.0% (4 in 408) in goats and 1.0% (5 in 486) in sheep. Among the samples 19.3% were seropositive for at least one of the three NZDs, 2.5% showed specific antibodies against at least two pathogens and 4 cattle (0.8%) were seropositive for all three pathogens. In cattle, adults over 3-years old and females presented a higher antibody seroprevalence for the three diseases, in congruence with putative exposure risk.This study confirms the circulation of these three zoonotic pathogens in Guinea and highlights the need for implementing a syndromic surveillance of ruminant abortions by the Guinean veterinary authorities as well as for the screening of the human population at risk  in a One Health perspective.The online version contains supplementary material available at 10.1186/s12917-022-03159-x. More than two-third of the newly emerging infectious agents affecting humans are zoonotic in origin . NeglectBrucella. Brucella abortus affects cattle, Brucella melitensis is found in small ruminants and Brucella suis in pigs. Transmission between animals mainly occurs by ingestion of infected birth fluids but also through cuts in the skin, or through mucous membranes. Brucella can cause fever, arthritis and neurological symptoms in humans [Brucellosis is caused by gram-negative bacteria from the genus n humans , 5. Brucn humans . In Afrin humans , 7 with n humans .Coxiella burnetii [C. burnetii is shed in milk, urine and feces but most importantly, during parturition in amniotic fluids and placenta. Transmission can also occur by spore inhalation or by tick vectors between animals. In humans Q fever is mainly asymptomatic or provokes self-limited illness with flu-like symptoms and transient hepatitis and pneumonia. Rare cases progress into chronic infection with endocarditis or vascular infection which can be lethal [Q Fever is caused by the intracellular gram-negative bacterium burnetii . A varieburnetii . Q Feverburnetii . In Westburnetii . C. burne lethal . In Afrie lethal . It can e lethal , 15.Bunyavirales, family Phenuiviridae, genus Phlebovirus [Aedes spp. and Culex spp. [Rift Valley fever (RVF) is a viral infection caused by a negative strand RNA virus belonging to the order ebovirus . Rift Vaebovirus , 18. RVFlex spp. , 20. Sinlex spp. , 22. A slex spp. . The higlex spp. .C. burnetii circulation with a mean seroprevalence of 2.4% (range 0.8–10.5%) in humans and of 8.0% (range 3.2–18.7%) in livestock [Guinea is situated in an intermediate position in Africa, connecting different ecosystems: a rather dry Sahelian area in the North in contact with Senegal and Mali, a wet forest area in the South in contact with Ivory Coast, Liberia and Sierra Leone, and the Atlantic littoral in the West. Only limited studies have been performed to evaluate the prevalence of brucellosis, Q fever and RVF in the country. The first recorded data of bovine brucellosis showed an overall seroprevalence of 6.9% range 0–27%) depending on the geographical location . Since t% dependiivestock . RVFV ciivestock . Anotherivestock . More reivestock .Up to now, all seroprevalence studies in Guinea focused only on cattle and the circulation of all three pathogens was never investigated simultaneously on the same animals during the same period. The purpose of the present study was to generate updated epidemiological data in the three diseases on domestic ruminants  in Guinea. Information on seroprevalence and risk factors associated with these three zoonoses will be useful to evaluate the associated risks on human health and animal production and to define suitable prevention and control programs in Guinea.Brucella spp., C. burnetii and Rift Valley fever virus (RVFV) using multi-species ELISA kits (IDVet). The overall seroprevalence was significantly different between the three diseases: Brucella spp. is 3.9%  2.9—5.1); Q fever is 9.1% (95% CI 7.7—10.8), RVF is 6.3% (95% CI 5.1—7.7) (Table S2). Interestingly, seroprevalence in cattle for the three pathogens is 5 to 20-fold higher than in small ruminants (goat and sheep)  from 17 prefectures of Guinea , 0.4% (95% CI 0.05—1.5) and 0% (95% CI 0—0.9) respectively , 4.4% (95% IC 2.6—6.9) and 2.3% (95% IC 1.1—4.0) in cattle, goats and sheep, respectively , 1.0% (95% CI 0.3—2.5) and 1.0% (95% CI 0.3—2.4), respectively  were found seropositive for the three pathogens  were seropositive for one of the three zoonoses, and 34 of them (2.5%) showed specific antibodies for more than one zoonotic agent: 9 cattle and 1 sheep were seropositive for both This study provides, for the first time for Guinea, seroprevalence data on three major NZDs, i.e. brucellosis, Q fever and Rift Valley fever (RVF), conducted in a short-time period (2017 to mid-2019) on the same 1357 ruminants: 463 cattle, 408 goats and 486 sheep. Such an extensive comparison of the three NZDs in three different ruminant species has rarely been done in West Africa , 33. ConIn addition, seroprevalence was generally 5 to 20-fold higher in cattle than in goats and sheep. An obvious explanation is that cattle live longer than small ruminants and have more time/opportunity for infections, which then leads to long detectable antibodies. This is evident from the mean age of the available samples in animals: cattle , goats  and sheep  Table .C. burnetii present in amniotic fluids and placenta during parturition and the spore-like form highly resistant to heat and dry once in the environment can explain the rapid dissemination by wind for long distances.While Q fever was previously considered a rare and regionally restricted tropical disease in Africa (11), our results show that it circulates in many prefectures in Guinea. This is in agreement with a previous report of seroprevalence in cattle in Upper Guinea (6.6 ± 1.3%), Forest Guinea (7.8 ± 1.7%), Central Guinea (9.1 ± 1.0%) and Maritime Guinea (7.5 ± 1.3%) . In the In contrast, brucellosis is more prevalent in the West  than in Central-East Guinea. The reasons for this geographical discrepancy remain to be clarified but could be linked to the fact that the bacteria better survive in an environment with cool moist conditions as in West Coastal Guinea. Our results are consistent with the finding of two earlier studies: one from the 1980s, which reported seroprevalence in Kindia (7.4%), Coyah 13.3%) and Forécariah (20%), but not in Dalaba or Kankan  and a se.3% and FRegarding RVF in cattle, we found an overall seroprevalence of 16.4% (0 to 35.0%) that was significantly higher than in previous studies (0 to 6.8%) , 29, 30.C. burnetii and Brucella sp. infections in cattle. In the prefectures of Dabola, Dalaba, Faranah, Kouroussa and Mandiana, where Q fever prevalence is higher (> 20%), brucellosis prevalence is null. In Kindia and Gaoual prefectures, where brucellosis prevalence is higher (> 40%), Q fever prevalence is null or low. The reason for this potential interference resulting in bi-directional antagonistic effects should be more precisely investigated. Specific immune cross-protection is improbable due to the genetic distance between both bacteria. Competition for the same ecological niche in term of infection or transmission has to be explored. It must be also noted that both bacteria  are able to survive for several months in the environment [Aedes sp. and Culex sp.), it is noteworthy that Dalaba has the highest rate of Q fever.An interesting finding of the present study is that several animals were found seropositive for more than one of the three NZDs suggesting that they have been successively or co-infected by the corresponding pathogens. Globally, the number of animals seropositive for more than one disease is statistically coherent based on the seroprevalence of each infection suggesting minimal influence between infections Table . Howeverhigher > 0%, Q fevB. abortus rather than B. melitensis in Guinea [B. abortus [B. melitensis has been associated with brucellosis epidemics in small ruminants mainly in Northern, Eastern and Central African countries [Brucella spp. are circulating among the ruminants in Guinea.Up to now, most studies on brucellosis seroprevalence in Africa have been focused in cattle only –27, 35. n Guinea , 36, 37.ountries . In thisAedes ochraceus, Aedes mcintoshi, and Mansonia uniformis have feeding preference for goats, followed by cattle, donkeys, sheep, and humans [RVF seroprevalence studies in African ruminants have delivered contradictory results . Most ofd humans . Entomold humans , 47. Thed humans , 24. In Consensus exists that Q fever seroprevalence is higher in goat and sheep is than in cattle . This isC. burnetii and Brucella spp. are found in amniotic fluid and placenta. Finally, female can be infected by Brucella spp. several times during life from parturition time with boost at each new birth. This explains that cattle seroprevalence is lower under 3 years old and steadily increase with age [Our study indicates that in cattle, females have higher risk to be infected than males by Q fever , RVF (threefold) and Brucellosis (fivefold). Similar results were previously reported across Africa , 36 and with age . In addiTritrichomonas foetus, Camphylobacter spp. or Bovine Viral Diarrhea virus. Multiplex serological (Luminex) or genetic  tools encompassing the main suspected aetiological agents should be developed in this perspective. Moreover, given the high bovine seroprevalence level of brucellosis, Q fever and RFV in some Guinean prefectures, it will be important to set up a One Health approach assessing the incidence of these zoonotic diseases in human and investigating the routes of transmission, in particular in people having close contacts with cattle such as breeders, veterinarians and/or slaughterers.Although targeting only asymptomatic animals, the present study highlights the circulation of three abortive animal diseases in Guinea with occurrence of simultaneous or consecutive co-infection. This invites further studies to investigate the mechanisms of interference between infections by the longitudinal follow up of selected farms from different ecological area in Guinea. Our results also invites the veterinary services to reinforce syndromic surveillance of ruminant abortions. Collection of vaginal swabs, placenta and/or abortive fetus could allow to better identify the infectious agents of these three diseases and to differentiate them from other pathogens provoking abortion such as A total of 1357 blood samples  from 17 prefectures of Guinea were included in this study  and centrifuged for 10 min at 3 000 RPM as rapidly as possible in the field to avoid hemolysis. Sera were stored frozen  until the serological tests were carried out.Brucella sp., C. burnetii and RVFV: ID Screen Brucellosis Serum Indirect Multi-Species, ID Screen Q-Fever Indirect Multi-Species and ID Screen Rift Valley Fever Competition Multi-Species, respectively. ELISA tests were performed and validated according to the manufacturer’s protocols and the test results were interpreted as positive, doubtful or negative. All sera giving positive or doubtful results in the first ELISA were retested a second time using the same ELISA kit. Only samples confirmed positive by the second test were considered positive, all the others were considered negative.Before serological analysis all sera were inactivated at 56 °C for 30 min. Sera were run in three different multi-species Enzyme-Linked Immunosorbent Assays (ELISA) from IDVet for detection of specific antibodies against ◦C with 5% CO2 for 1 h. Next, the virus/serum mix was inoculated onto VeroE6 cell monolayers in 12-well-plates (5 × 105 cells per well). One hour after adsorption at 37 °C, 2 mL of diluted carboxymethylcellulose sodium salt with DMEM supplemented with 5% FBS (v/v) were added to each well and plates were incubated at 37 °C in 5% CO2 for 5 days. The plaque forming units (pfu) were then revealed by 1% (w/v) crystal violet and the number of pfu/mL was calculated for each serum. The neutralizing Ab titers were established as the last dilution which inhibited 50% of the foci number per well compared to virus-only control titration.Neutralizing antibodies (Ab) against RVFV were measured as previously described in BSL3 conditions , 61. SerData were analyzed with the GraphPad Prism 6.0 software. The unpaired non-parametric Mann–Whitney test was used to compare female and male seroprevalence for each disease and species. The non-parametric Kruskal–Wallis test followed by Dunn’s correction was used to compare the seroprevalence among age groups and prefectures 2 by 2 for each diseases and species.Additional file 1.


As insecticide-treated nets (ITNs) wear out and are disposed, some household members are prioritized to use remaining ITNs. This study assessed how nets are allocated within households to individuals of different age categories as ITNs are lost or damaged and as new ITNs are obtained. The study also explored how ITN allocation affects ITN durability.A cross-sectional household survey and ITN durability study was conducted among 2,875 households across Tanzania to determine the proportion of nets that remain protective (serviceable) 22 months after net distribution aiming for universal coverage. Allocation of study nets within houses, and re-allocation of ITNs when new universal replacement campaign (URC) nets arrived in study households in Musoma District, was also assessed.Some 57.0% (95% CI 53.9–60.1%) of households had sufficient ITNs for every household member, while 84.4% (95% CI 82.4–86.4%) of the population had access to an ITN within their household (assuming 1 net covers every 2 members). In households with sufficient nets, 77.5% of members slept under ITNs. In households without sufficient nets, pregnant women (54.6%), children < 5 years (45.8%) and adults (42.1%) were prioritized, with fewer school-age children 5–14 years (35.9%), youths 15–24 years (28.1%) and seniors > 65 years (32.6%) sleeping under ITNs. Crowding (Users determined the useful life of ITNs and prioritized pregnant women and children < 5 to serviceable ITNs. When household net access declines, users adjust by crowding under remaining nets, which further reduces ITN lifespan. School-age children that commonly harbour gametocytes that mediate malaria transmission are compelled to sleep under unserviceable nets, crowd under nets or remain uncovered. However, they were accommodated by the arrival of new nets. More frequent ITN delivery through the school net programme in combination with mass distribution campaigns is essential to maximize ITN effectiveness.The online version contains supplementary material available at 10.1186/s12936-021-03686-2. Insecticide-treated nets (ITNs) are impactful in the fight against malaria in sub-Saharan Africa . In TanzEffective malaria protection by ITNs is achieved when at least 80% of household members have access to, and sleep under ITNs . The WorThere are several factors that affect ITN access and use, including household size , user chIn 2015, a cross-sectional household survey was conducted in 2,875 households across eight districts. The survey was conducted between October and December just before the short rainy season when malaria transmission is usually low. The households randomly received one type of ITN from a pool of 3 products (referred hereafter as study nets): Olyset®, NetProtect®, PermaNet®, to cover every sleeping space identified during enrolment in 2013. Participating households were also geo-referenced using global positioning systems (GPS) during enrolment to aid identification during follow-up visits. Study nets were identifiable by their colour (white) and with a durable waterproof label to allow longitudinal follow-up. The average number of sleeping spaces per household among the study population was 3.1 and each household received an average of 3 study nets. Study-net dimensions were of double size (190 cm × 180 cm × 150 cm) assumed to fit 2 people under each net, similar to those distributed by the NMCP .Data were generated as part of a longitudinal ITN durability study with three data collections  , 32 but Over time, nets become torn with repeated use. While the inclusion of pyrethroid insecticides helps to prevent mosquitoes entering nets to some extent , the morAn in-depth assessment of some of the Roll Back Malaria Monitoring and Evaluation Reference Group (MERG) indicators , 38 as wData analysis was carried out using statistical software package STATA 14.1 . Survey weights were used to compensate for unequal sampling units, adjust for non-response, and a multi-level modelling approach. Net use and the proportion of serviceable and unserviceable study nets by user age category, among houses with and without enough nets for every 2 members, are presented as frequencies and percentages. Statistical analysis of the effect of crowding (more than 2 people sleeping under a net) on net serviceability was done using logistic regression models with crowding as the main exposure. Other predictor variables specified a priori were user characteristics , SES and net product. A forward-selection procedure was applied for modelling and the selection was based on change in main exposure effect estimate (mean square error). The procedure involved three main steps: (a) descriptive analysis and preliminary investigations for association between variables while paying attention to the sizes of effects as well as two-sided p-values at 95% significance level; (b) variables selection: from prior knowledge, age and gender were considered as forced variables in the model. Then, one variable at a time from a list of candidate variables obtained from univariate analysis was included in the model with and without adjustment of forced variables to understand the effect of forced variables. The choice of the ‘best’ predictor to be included in the model was then decided based on the change in exposure effect estimate. Each time a new variable was added in the model, evidence of confounding and multicollinearity was assessed by comparing the effect estimates and standard errors between the ‘univariate’ and ‘multivariate’ models estimates; and, (c) multivariable models were fitted by adding explanatory variables that were removed from the models in step (b), one at a time to explore their effect when added to the model in presence of other variables in the model. Variables that resulted in positive changes in the mean square error were then included in the model. The process was repeated until all variables that provided precise estimates of exposure variables were selected.A total of 3096 households were re-visited from the 8 participating study districts of which 2875 were interviewed yielding a response rate of 92.9%. Of the 221 households not interviewed, 110 households withdrew from the study as they had moved away from the study village, 70 householders were temporarily unavailable (in farms), while 41 households withdrew consent to continue to participate. Mosquito nets were found in 2801 (97.4%) households of which 1668 (58.0%) had study nets, 1126 (39.2%) had both study and non-study nets, and 7 households (0.2%) had non-study nets. Overall, 9178 mosquito nets were found, of which 5899 were in households with sufficient ITNs and 3288 in households without sufficient ITNs. Of these mosquito nets, 6938 (75.6%) were identified as study nets and 2249 (24.5%) as non-study nets since they were obtained from other sources. Of the non-study nets, 712 (31.7%) were identified as ITNs based on their product label. Therefore, a total of 7650 ITNs (study and non-study) were identified and included in the analyses presented.In 2013, as part of the study design, 100% of sleeping spaces were covered by study nets, and this fell to 42.6% of sleeping spaces covered by study nets after 22 months. Including study nets and non-study ITNs, 57% (95% CI: 53.9–60.1%) of the participating households still had sufficient ITNs, i.e., one ITN for every 2 household members assuming each ITN is used by 2 people. Eighty-four per cent (95% CI: 82.4–86.4%) of the population living in the participating households had access to an ITN, assuming each ITN was used by 2 people, and 53.2% (95% CI: 52.4–54.0%) of those with access used an ITN the previous night  contributed the lowest percentage of ITN users . For children < 5, 82.9% slept under an ITN, 5.4% higher than the household average of 77.5%. In houses without enough nets, 45.8% of children < 5 slept under ITNs, which is 8.4% higher than the household average use of 37.4%. A slightly smaller proportion of children 5–14 years slept under ITNs compared to the household average in both houses with enough nets (75.7 vs 77.5%) and in households without enough nets (35.9 vs 37.4%). Youths were also less likely to be prioritized to ITNs in houses with enough nets (5% lower than household average) and this was more pronounced in houses without enough nets (9.3% lower than household average). Seniors were less likely to be prioritized to ITN use in houses without enough ITNs, with only 32.6% of them sleeping under nets which was 4.8% lower than the household average, although this was not seen in houses with sufficient ITNs.In households with enough nets, 91.1% of pregnant women slept under ITNs, 13.6% higher than the household average of 77.5% use. In houses without enough nets, a 17%-point increase in net use among pregnant women was observed when compared to the average household use  had ITNs that were used the previous night. Of the 3,288 mosquito nets found in households without enough ITNs, 25.1% 95% CI: 23.0–27.3%) were used by 3 sleepers, while 8.8% (95% CI: 8.0–9.7%) of the 5,899 nets found in households with enough ITNs were used by 3 or more people. The proportion of 3 or more household members sleeping under one net was higher in households without enough ITNs (62.1% (95% CI: 60.7–63.6%) compared to those with enough ITNs [30.5% (95% CI: 29.2–31.7%) (Table .0–27.3% Holes were counted in 4783 (68.9%) of the 6938 study nets, 22 months after distribution. Of these, 3735 78.1%) nets were still serviceable while 1,048 (21.9%) were unserviceable. Only 3622 (75.7%) of the 4783 ITNs assessed for physical damage were used the previous night. Furthermore, 847 (80.8%) of unserviceable nets and 2775 (74.3%) of serviceable nets were used last night. Prioritization of serviceable nets was also observed. On average, 32.6% people slept under serviceable ITNs the previous night whereby around 7% more pregnant women (40.5%), adults (39.2%), seniors (39.3%), and 5% fewer children 5–14 (27.8%), and 6% fewer youths 15–24 (26.6%) slept under a serviceable ITN  in the univariate analysis. The odds of NetProtect® nets being serviceable was two times the odds of Olyset® nets 2.08 (95% CI 1.68–2.58), p < 0.001. ITNs used by children (5-14 years) had lower odds of being serviceable compared to those used by under-fives 0.72 (95% CI 0.56–0.93), p < 0.001. Controlling for net product and user characteristics , crowding was significantly associated with unserviceable ITNs (p < 0.001). Compared to one person under a net, having two people under the net reduced the odds of serviceability to OR 0.75 (95% CI: 0.60–0.83) and having three people under the net further reduced the odds of serviceability to OR 0.50 (95% CI: 0.40–0.63).Results of univariable and multivariable analyses exploring the consequences of net allocation on ITN serviceability are presented in Table A total of 398 households were visited in Musoma district by the study team in 2015, where 7 households were found with no nets. The average number of sleeping spaces per household was found to be 3.3 and the average number of people per household was 6.1. Forty-four per cent (95% CI: 38.8–48.8%) of households had at least one URC net with an average of 1.4 URC nets per household. Ten per cent (95% CI: 9.2–12.6%) of the households had enough URC nets; 23.6% (95% CI 16.7–30.6%) of the population in those households had access to a URC; 27.7% (95% CI 25.9–29.5%) of the population used a URC net the night before the survey  the previous night in comparison to 13.0% under URC nets and 12.0% under nets acquired from other sources , the number of serviceable nets in a household decreases. Eighty per cent of household members were observed to sleep under a net when the person: net ratio was 3:1 and this decreased to 50% of the population using a net when 4 or more people slept under a single net, with the remaining 50% being left uncovered . While tPhysical degradation of the net products was also observed to vary by product after 22 months of ownership. NetProtect® was two times more likely to be serviceable when compared to Olyset® in this setting. When compared to PermaNet®, Olyset® nets have been observed to have more holes in both Mozambique , Zambia . [Population access was 84.4% just prior to the URC campaign in the study population, with the exception of Musoma district which had already received campaign nets, and which in addition to study nets, increased access to 94.3%. Unfortunately, despite the URC that was conducted August 2015 to January 2017, none of the participating districts recorded an increase in population access according to the TMIS  that was.  in both . , and wil. , which w. . The sch. –57. That. [Increasing access to nets within a household increases net use, which in turn will eliminate inequalities between age and gender . Contrar. , where i. , 59 and . , 60), whThe study distributed one ITN for every sleeping space identified during enrolment instead of using the recommended practice of one ITN for every 2 household members. While this distribution mechanism may have prevented distribution of excess ITNs to household members without unique sleeping spaces, it biased household and population access to ITNs to higher levels than would be achieved by national campaigns from enrolment.It is also important to note that the URC coincided with the general election season in Tanzania. This may have contributed to the lack of extensive distribution of ITNs in Musoma district and/or the delayed distribution of ITNs to populations prior to this study in order to prevent affiliation with any political party rally coinciding with a distribution date.There is also a challenge in the definition and measurement of population access in assuming each ITN is used by 2 people. For example, if a 25-years-old woman is living with her uncle and they have only one net, in principle as per MERG indicators for measuring household mosquito net distribution, population access is complete. However, in practice, these two people are unlikely to sleep under the same net, leaving one household member uncovered and population access incomplete. Therefore, this was a challenge while assessing population access that could not be changed or controlled.While even torn nets still offer chemical protection against mosquitoes , 62, incTwenty-two months post-ITN distribution, over 50% of sleeping spaces did not have access to a study net, despite complete coverage at baseline. However, the percentage of the population with access to ITNs was above the target of 80% while 57% of households had enough ITNs. The URC mass campaign helped to further maintain universal access to ITNs in Musoma district. These findings indicate that households hold on to their ITNs despite the arrival of new ones. Crowding under ITNs was associated with lower ITN serviceability most likely due to physical stress on the ITN fabric that causes physical damage to occur faster, thereby reducing the serviceable life of the net. When households have enough nets, around 80% of members from all age categories have access to a net. However, when there are insufficient nets, children (5–15 years) and youths (15–24 years) were least likely to use any ITN or have access to a serviceable ITN. This is of significant biological importance since school-aged children carry gametocytes that cause transmission of malaria from humans to mosquitoes and maintain malaria transmission. There is a need to refine delivery strategies to ensure households, including larger households, receive sufficient nets to cover all sleeping spaces. More frequent and more informed ITN distribution through keep-up strategies such as the school-net programme is essential to address these coverage inequalities and ensure continued protection against malaria transmission for all household members.Additional file 1. Additional file 2: Table S1. Coverage of nets in Musoma district by net source.


Gracilaria fisheri. The dried seaweed sample was digested using three different methods under various conditions using enzymes (cellulase and protease), 30% H2O2, and a combination of enzymes and 30% H2O2. The method possessing the best digestion efficiency and polymer recovery rate of MPs was selected, and its effect on spiked plastic polymer integrity was analyzed by Raman spectroscopy. As a result, the enzymatic method rendered moderate digestion efficiency (59.3–63.7%) and high polymer recovery rate (94.7–98.9%). The oxidative method using 30% H2O2 showed high digestion efficiency (93.0–96.3%) and high polymer recovery rate (>98%). The combination method was the most effective method in terms of digestion efficiency, polymer recovery rate, and expenditure of digestion time. The method also showed no chemical changes in the spiked plastic polymers  after the digestion process. All the spiked plastic polymers were identifiable using Raman spectroscopy.Currently, research on the accumulation of microplastics (MPs) in the marine food web is being highlighted. An accurate and reliable digestion method to extract and isolate MPs from complex food matrices has seldom been validated. This study aimed to compare the efficacy of MP isolation among enzymatic-, oxidative-, and the combination of two digestion methods on red seaweed,  Annual plastic production reached 381 million tonnes in 2015, which was around 200-fold higher than the total amount of plastic produced in the 1950s . The abuMPs can be accumulated in a wide variety of organisms ,10,11 anOver the last few years, attempts to detect and analyze MPs in food items have been made to assess and manage the risks associated with MP exposure. In MP analysis, the digestion of food matrices is crucial to the dissolution of organic matter and the isolation of synthetic polymers from the food samples. A lack of standard digestion methods for MP analysis results in the difficulty to compare the effectiveness of digestion performance. The quality of different digestion methods can be assessed through the ability to reduce the matrix complexity, i.e., digestion efficiency and to prevent plastic polymer degradation, which is usually demonstrated as polymer recovery rate .3) [3 [2O2) [Gracilaria fisheri (G. fisheri) has not yet been conducted.To isolate MPs from marine animals, previous research applied different digestion methods including acid ,22, alka3) . Meanwhi3) [3  and a mo[3 [2O2) . To the 3 obtained a higher digestion efficiency than potassium hydroxide (KOH) and H2O2 in prawn and mussel, respectively [2O2) was more efficient than the use of alkaline solution to digest biogenic matter in marine sediment [2O2 concentration and temperature caused a destructive effect, especially on synthetic PA [4), obtained high digestion efficiency (>97%) and did not cause any destructive effect on PS, PE, PVC, nylon, and polyester [Among chemical digestions, HNOectively ,32. Howeectively ,33. For ectively . Howeverectively . Oxidatisediment , especiasediment . Howeverhetic PA . Enzymathetic PA . Howeverhetic PA . For comolyester .G. fisheri, an edible red seaweed naturally distributed in the shoreline area, is widely used as an ingredient in commercial food products [G. fisheri in treating cholangiocarcinoma in human and white spot syndrome in shrimp are obtained from sulfated galactan, which is a polysaccharide found in the cell walls of marine algae [G. fisheri and other varieties of seaweed are becoming important indicators for assessing MP contamination in the marine environment as they can potentially trap MPs from the surrounding water [G. fisheri is still a challenging issue in the field of MP research.products . Moreoveproducts  and as aproducts . The rolne algae ,39. The ne algae , while tne algae . G. fishng water ,43. OwinG. fisheri. Three digestion methods, including enzymatic-, oxidative- and combination of enzymatic and oxidative methods, were studied for the digestion efficiency, polymer recovery rate, and microscopic observation under a stereomicroscope. In addition, the chemical composition of each spiked plastic polymer after the digestion process using an optimal method in suitable conditions was further analyzed by using Raman spectroscopy.This study aimed to compare the effectiveness of different digestion methods in various conditions for isolating MPs from dried G. fisheri collected from the coast of the Gulf of Thailand was resized into 2–5 mm lengths. The sample was stored in a sealed aluminium foil bag prior to use. The analysis of the moisture content of dried G. fisheri was carried out according to the Association of Official Analytical Chemists (AOAC) method no 930.04 [Dried o 930.04 .MP contamination control was applied throughout the entire analytical procedure  to avoid contamination from workers, environment, and equipment. All work surfaces were thoroughly cleaned using 70% ethanol to prevent contamination . A negatG. fisheri  was transferred to a 250 mL Erlenmeyer flask containing 25 mL of sodium phosphate buffer saline . The flask was incubated at 50 °C, 90 rpm for 30 min prior to simultaneous addition of cellulase  and protease . The flask was then further incubated at 50 °C, 90 rpm for 30 h. The concentration of cellulase used was varied as follows: 1%, 3%, and 5% (v/v), while the concentration of protease used was 5% (v/v) in all enzymatic conditions.Dried 2O2  at 1:50 (OD50) and 1:100 (OD100) were studied. The 30% H2O2 was added to each 0.25 g (DW) of dried G. fisheri. Then, the mixture was incubated at 60 °C, 90 rpm for 96 h. Two different weight by volume ratios of sample to 30% HG. fisheri was digested by using the enzymatic and oxidative methods in sequential order. Briefly, 0.25 g (DW) of dried G. fisheri was digested by using 1% (v/v) cellulase and 5% (v/v) protease at 50 °C, 90 rpm for 2 h. Then, the enzyme was inactivated at 85 °C for 10 min. Subsequently, 30% H2O2 was added with the sample to H2O2 ratio of 1:100 and was incubated at 60 °C, 90 rpm for 36 h.g for 10 min. The supernatants were collected and stored at 4 °C for subsequent 3,5-dinitrosalicylic acid (DNS) and 2,4,6-trinitrobenzene sulfonic acid (TNBS) assays.Enzymatic digestates (600 μL) collected at different time points (0 to 30 h) were placed in a water bath at 85 °C for 10 min to inactivate enzymes. Then, the solutions were rapidly cooled on ice for 5 min prior to centrifugation at 1000× The DNS assay described by Miller (1959) was performed to monitor the enzymatic hydrolysis of carbohydrates through measuring the released reducing sugars  The DNS The TNBS assay described by Benjakul and Morrissey (1997) was performed to monitor the enzymatic hydrolysis of protein through measuring the released α-amino acids . The 0.0g for 10 min. The supernatant was then collected and filtered through a 2.5 μm Whatman™ filter paper  using a vacuum pump  with a pressure of approximately 20 to 40 kPa. The filter paper with retained digestate from the seaweed sample was dried to constant weight at 70 °C. The weight of the digested sample on the filter paper was used to calculate the digestion efficiency using the following equation [i = Initial weight of seaweed sample; Wa = Weight of dry filter paper after filtration; Wb = Weight of dry filter paper before filtration.After digestion, the sample was centrifuged at 1000× equation .(1)Digei = Initial weight of spiked plastic particles; Wa = Weight of dry filter paper after filtration; Wb = Weight of dry filter paper before filtration.For the polymer recovery rate, five types of plastic polymers, including medium-density polyethylene (PE) , polypropylene (PP) , polystyrene (PS) , polyethylene terephthalate (PET) , and polyvinyl chloride (PVC) , were analyzed as the control spikes in parallel with all the digestion conditions. The positive control contained 2 mg of each type of plastic polymer and 1 pellet of PET. The total weight of the five spiked plastic particles on the filter paper was used to calculate the polymer recovery rate using the following equation .(2)PolyTo evaluate the effect of each digestion method, the digestion efficiency and polymer recovery rate obtained from each digesting condition were substracted with those obtained from negative controls. To determine the effect of the digestion procedures on the plastic polymers, the obtained digestates and spiked plastic particles retained on the surface of the dried filter paper were visualized by using a stereomicroscope  at 10× and 40× objectives with ZEN 3.0 Blue edition software.−1, with an acquisition time of 20 s repeated three times. The equipment was calibrated with silicon wafer prior to use. The Raman system was operated using LabSpec6 software. After the detection process, the spectra of the spiked plastic polymers were compared with the reference plastic spectra in the KnowItAll spectral library .In this study, the spiked plastic particles that underwent the combination digestion method were characterized by Raman spectroscopy to investigate the effect of the digestion process on the integrity of each polymer. The Raman spectroscopy system  was equipped with 500 μm confocal hole, 100 μm slit, and 600 grooves/mm grating. Then 785 nm radiation of laser and a 50× objective were applied. Raman spectra were recorded in the wavenumber range of 200–3500 cmp < 0.05 was considered to be statistically significant. The data were analyzed using the IBM SPSS Statistics for Windows version 26.0 software .All experiments were conducted at least in triplicate. The results were expressed as mean ± standard deviation when appropriate. Statistical analysis was performed using Analysis of Variance (ANOVA) with Duncan’s Multiple Range test. The v/v) in combination with 5%, v/v, protease were tested. Enzymatic digestion was performed at 50 °C with shaking. The DNS and TNBS assays were applied to monitor the hydrolysis of G. fisheri by cellulase and protease through the analysis of reducing sugars and free amino groups, respectively.Three concentrations of cellulase  with increasing cellulase concentration, and the rates obtained with 3% and 5% (v/v) cellulase were 2-fold and 3-fold higher than the rate obtained with 1% (v/v) cellulase, respectively. After 15 min of digestion, the cellulose hydrolysis reached a plateau where no significant increase (p > 0.05) in reducing sugar concentration over time was observed. At the end of observation at 6 h, reducing sugar concentrations obtained with 1%, 3%, and 5% (v/v) cellulase  were significantly different (p < 0.05).Based on the released reducing sugar, a similar hydrolysis profile of nditions . OverallG. fisheri protein based on the release of free amino acids  when the digestion was prolonged from 4 h to 6 h; hence, this suggested that the hydrolysis reaction was likely to reach a plateau after 6 h of digestion. At the end of observation at 6 h, the free amino acid concentrations obtained with 1%, 3%, and 5% (v/v) cellulase combined with 5% (v/v) protease were 2.60 mg/mL, 2.97 mg/mL, and 3.13 mg/mL, respectively. It was noted that the higher cellulase concentration seemed to promote the hydrolysis activity of protease.Similar to the results of cellulose hydrolysis, all tested enzymatic conditions resulted in a resemble hydrolysis profile of no acids . As a whG. fisheri are shown in v/v) cellulase and 5% (v/v) protease. This finding was in agreement with the results obtained from DNS and TNBS assays, which showed that an increased cellulase concentration had a more pronounced effect on the degradation of G. fisheri. Conversely, the digestion with 5% (v/v) cellulase and 5% (v/v) protease resulted in the lowest recovery rate of the spiked plastic particles (94.7%), while the use of 1% (v/v) cellulase and 5% (v/v) protease rendered the highest polymer recovery rate (98.9%).The digestion efficiency and polymer recovery rate observed after 30 h for enzymatic digestion of 2O2 ratios, including OD50 and OD100, were tested to determine the digestion efficiency over 96 h of digestion. As shown in G. fisheri was digested almost completely at 6 h of oxidative digestion at which the digestion efficiencies observed for OD50 and OD100 were about 93.0% and 93.4%, respectively. The efficiencies of OD50 and OD100 reached over 95% after 48 h and 24 h of digestion, respectively, whereas the increase in digestion time from 48 h to 72 h did not result in any significant improvement (p > 0.05) in digestion efficiency. Surprisingly, the efficiencies of both OD50 and OD100 significantly decreased (p < 0.05) when extending the digestion time to 96 h.For the oxidative method, two samples at 30% H2O2 and digestion duration on the rate of polymer recovery, and oxidative digestion with all-time intervals conferred high recovery rates above the value of 95%, which was set by Karami et al. (2017) [For the polymer recovery rate of the oxidative method, there was no significant effect of 30% H. (2017) . HoweverA noticeable amount of retained seaweed on the filter paper was clearly observed for OD50 treatment. This might interfere with the further analysis of MP visual identification under the stereomicroscope . For botv/v) cellulase and 5% (v/v) was performed in the first step, followed by the 36 h of oxidative digestion using the ratio of sample to 30% H2O2 of 1:100. The enzymatic digestion condition was selected based on the best polymer recovery rate (98.9%), while the oxidative digestion condition was selected based on the best digestion efficiency and microscopic observation results. As shown in G. fisheri.The combination method involves two steps, of which the 2 h of enzymatic digestion using 1%  combined with 5% (v/v) protease led to the improvement of digestion efficiency from 59.3% to 63.7% and the increase of released reducing sugar. The improvement of cellulose hydrolysis was correlated to the increase of cellulase concentration as reported by Kumari et al. (2020) [G. fisheri also contains hemicellulose and lignin [Enzymatic approaches are known to possess several advantages over chemical methods such as mild process conditions and high specificity. However, the major limitations of the enzymatic method are the cost and time required . In the . (2020) . However. (2020) . Apart fd lignin  which arThe polymer recovery rate obtained using the enzymatic method was considered a high recovery rate, categorized based on the value, between 95% to 105%, which was set by Karami et al. (2017) . In this2O2 is one of the most versatile oxidative and bleaching agents. H2O2 can digest a wide range of carbohydrates, proteins, and lipids as the result of the high digestion efficiency and polymer recovery rate as observed in many food materials [2O2 at a high concentration (30–35%) was used to digest mussel and fish samples for MP analysis [2O2 ratio used in the digestion [2O2 at 50 °C and reported a decrease of digestion efficiency of about 10.2% from 1 h to 6 h, but gave no explanation of this finding [2O2 in different food matrices. For example, the study of Prata et al. (2019) showed an improvement in digestion efficiency and shortened the digestion time by using 30% H2O2 with a catalyst (Fe(II)) [2O2 digestion are the need for high temperature and time consumption [Briefly, this study showed that 30% Haterials ,24. In panalysis ,11,21,24analysis . Howeverigestion . In this finding . Meanwhi(Fe(II)) . The limt al. 201 showed asumption , besidesation 30–% was usesumption .2O2 has pH value ≤ 3.5 [G. fisheri. Mintenig et al. (2017) and Hurley et al. (2018) also reported the effectivity of enzyme–oxidative combination for digesting the organic matter for MP analysis [For the combination method, the enzymatic digestion was performed prior to oxidative digestion to avoid the disruption of protease and cellulase activities which could be influenced by the changes in pH value. According to the chemical data, 30% Hue ≤ 3.5 , while tue ≤ 3.5 ,59. A prue ≤ 3.5 . The comanalysis ,61.−1. This range corresponded to C–H vibration [−1. This spectral range corresponded to aromatic bending vibration [G. fisheri seaweed and exhibited a high potential to be employed as the sample preparation method for MP analysis in other polysaccharide and protein-rich materials. Moreover, the time expenditure of this combination method was shorter than the previously reported protocol, which required 74 h for the digestion of nori seaweed [For polymer characterization, Raman spectroscopy is widely known due to its ability at a low limit of detection and wide spectral range . In thisibration . Given tibration . The absibration ,65. Over seaweed .G. fisheri. Significant effects of the enzymatic and oxidative methods, as well as the combination method on G. fisheri, were identified. The high polysaccharide content of algae is one of the most common obstacles in the isolation of MPs from the sample. The enzymatic method using cellulase and protease resulted in moderate digestion efficiency. Meanwhile, the oxidative approach afforded great compatibility with the seaweed digestion but it required a long digestion period. The combined use of enzymatic and oxidative digestion methods successfully destroyed the dried G. fisheri tissue and offered high MP retention. This method is suggested as a promising digestion protocol for MP analysis in G. fisheri and might be able to be extensively applied to other vegetal tissues for MP analysis.For research on MP analysis in marine organisms, diverse analytical digestion methods have been tailored for specific food matrices to achieve the removal of biogenic matter. This study highlighted the optimization of the digestion conditions for MP analysis in the red seaweed,


IntroductionTuberculous meningitis (TBM) brings significant morbidity and mortality worldwide. Hyponatremia has long been documented as a potentially grave metabolic result of TBM. The syndrome of inappropriate antidiuretic hormone (SIADH) secretion has been supposed to be accountable for the majority of cases of hyponatremia in TBM. Cerebral salt wasting syndrome (CSWS) is being progressively reported as a basis of hyponatremia in some of these cases. Differentiating CSWS from SIADH can be challenging but is vital because treatment of these two conditions is profoundly different.ObjectiveThe rationale of our study is to determine the frequency of hyponatremia and etiology in patients presenting with TBM in a tertiary care hospital in order to establish the local perspective as there is paucity of local data.MethodsA total of 160 hospitalized patients at a tertiary care hospital in Pakistan who fulfilled the inclusion criteria were enrolled in this study after informed consent. The study was conducted for six months at the department of neurology, Jinnah Postgraduate Medical Centre (JPMC), Karachi, Pakistan. Brief history was taken and demographic information was entered in the performa by researchers. The data was collected and analyzed on Statistical Package for Social Sciences (SPSS) version 18.0 . Demographic data were presented as simple descriptive statistics giving mean and standard deviation for age, height, weight, GCS , serum sodium and duration of symptoms. Frequencies and percentages were calculated for categorical variables like gender, hypertension, smoking status, T2DM (Type 2 Diabetes Mellitus), BMRC  stage, hyponatremia, SIADH and CSWS. Effect modifiers were controlled through stratification of age, gender, hypertension, smoking status, T2DM, BMRC stage and duration of symptoms to see the effect of these on the outcome variable (hyponatremia). Quantitative data were presented as simple descriptive statistics giving mean and standard deviation and qualitative variables were presented as frequency and percentages. Post stratification chi-square test was applied with a p-value of ≤0.05 taken as significant.ResultsIn our study, out of 160 patients with TBM, 40% (64) had hyponatremia. Moreover, 14.4% and 25.6% had SIADH and CSWS, respectively with 60% (96) of patients were male and 40% (64) were female. Mean age of patients in our study was 46.78±2.81 years. Whereas, mean duration of symptoms, serum sodium, GCS, height and weight in our study was 1.2±0.78 weeks, 128.65±7.52 mmol/L and 11.21±3.14%, 158±7.28 cm and 78.7±9.87 kg, respectively.ConclusionThis study concluded that the frequency of hyponatremia among patients of TBM was significant, consistent with previous studies. Privation of proper assessment and management can lead to grave and permanent neurological consequences, as well as death. Healthcare providers should be aware of the implication of sodium deregulation among patients of TBM and differentiate between the numerous therapeutic preferences in order to advocate safe and effective treatment. Tuberculosis (TB) still remains one of the foremost infectious sources of morbidity and mortality globally . AccordiHyponatremia is a common electrolyte disorder and is a common outcome in patients with acute cerebral insult . HyponatWe conducted a six-month cross-sectional study of consenting patients admitted with a diagnosis of TBM at the Department of Neurology, Jinnah Postgraduate Medical Center (JPMC), Karachi, Pakistan. In our study, inclusion criteria included all patients of either sex aged 18 to 60 years. Non-consenting patients were excluded from the study along with those patients with history of mania, bipolar effective disorder or posttraumatic stress, lung carcinoma, hypothyroidism or hyperthyroidism, Addison’s disease, tuberculosis, head trauma, multiple sclerosis, asthma, renal impairment, congestive heart failure, myocardial infarction, chronic obstructive pulmonary disease and chronic liver disease. A brief history of the duration of illness and demographic information was taken from each patient and confirmed by an attendant. Blood samples were drawn by the researchers and sent for the measurement of serum urea, electrolytes, thyroid function tests, short synacthen test and urinary sodium at the time of admission and transported to hospital standardized laboratory by proper labeling as well as the investigation requested. Patients were labeled as having hyponatremia, SIADH and CSWS as per operational definitions. The findings were noted in the performa along with other demographic and confounding variables like hypertension, smoking status, T2DM (type 2 diabetes mellitus), BMRC  stage I/II/III and duration of symptoms.Patients were diagnosed with TBM if presenting with any two or more of the clinical features in one week: Fever ≥ 99°F occurring about at least 6 hours/day for more than one week, headache dull in nature persisting on VAS  ≥ 6 for at least three hours/day per day for more than one week with 0 as no pain and 10 being most severe pain, vomiting of any amount at least three times per day for three consecutive days or history of contact with TB patient in family (living in the same house) or outside (outside the house) in last two years. Apart from clinical features presence of any one of the positive laboratory data was used to label TBM: positive acid-fast bacilli (AFB) smear on cerebrospinal fluid (CSF), positive AFB culture on CSF, CSF pleocytosis (20-500 lymphocytes per cubic mm) with increased CSF protein ≥ 100 mg/dl and decreased CSF glucose concentration <60% of corresponding plasma level checked at the same time as CSF examined. Patients having serum sodium level ≤135 mEq/L were labeled as having hyponatremia. Patients with hyponatremia was evaluated for the etiology of hyponatremia and patients was labeled as having SIADH or CSWS based on the volume status; Euvolemia (absence of supine heart rate more than 100 and systolic BP less than 100 mmHg) was labeled SIADH and hypovolemia (presence of supine heart rate more than 100 and systolic BP less than 100 mm Hg) was labeled as CSWS, plus having any two or more of the following: Plasma sodium concentration ≤135 mmol/L, Plasma osmolality ≤280 mOsmol/kg, Urine osmolality ≥100 mOsmol/kg, Urinary sodium concentration ≥30mmol/L, absence of clinical or biochemical features of adrenal and thyroid dysfunction or no history of diuretic use within the past three months. Patients were labeled as hypertensive if known hypertensive for more than two years on treatment and compliant (patients taking medicines regularly and having SBP ≤ 140 mmHg and ≤ 90 mmHg for more than six months). Patients were labeled as diabetic if known diabetic on treatment for more than two years on treatment and compliant (patients taking medicines regularly (daily) or having HbA1C ≥ 6.5%. Patient was labeled as smoker if a person smokes at least five cigarettes a day for at least one year.The data were collected and analyzed on Statistical Package for Social Sciences (SPSS) version 18.0 . Demographic data were presented as simple descriptive statistics giving mean and standard deviation for age, height, weight, GCS , serum sodium and duration of symptoms. Frequencies and percentages were calculated for categorical variables like gender, hypertension, smoking status, DM, BMRC stage, hyponatremia, SIADH and CSWS. Effect modifiers were controlled through stratification of age, gender, hypertension, smoking status, DM, BMRC  stage and duration of symptoms to see the effect of these on the outcome variable (hyponatremia). Quantitative data were presented as simple descriptive statistics giving mean and standard deviation and qualitative variables was presented as frequency and percentages. Post-stratification chi-square test was applied with a p-value of ≤0.05 taken as significant. We applied non-probability consecutive sampling. For sample size calculation we used OpenEpi, by taking frequency of SIADH 8.82% , margin A total of 160 patients diagnosed with TBM were included in this study according to the inclusion criteria. Out of 160 patients with TBM, 40% (64) had hyponatremia. It was also found that 14.4% (23) had SIADH and 25.6% (41) had CSWS. The age range of the patients was 26 years being minimum while the maximum age of the patients was 60 years. Mean age in our study was 46.78 years with a standard deviation of ±2.81. Whereas, mean duration of symptoms, serum sodium, GCS, height and weight in our study was 1.2±0.78 weeks, 128.65±7.52 mmol/L (range: 125-135) and 11.21±3.14, 158±7.28 cm and 78.7±9.87 kg respectively. In terms of gender distribution among cases of our study, 60% (96) of our participants were male and 40% (64) were female.Frequency distribution of age showed that out of 160 patients with TBM, 21.2% (34), 30% (48), 15.6% (25) and 33.1% (53) patients were in the age group 20-30 years, 31-40 years, 41-50 years and 51-60 years respectively as presented in Figure Frequency distribution of duration of symptoms showed that out of 160 patients with TBM, 50.6% (81) and 49.4% (79) had symptoms for less than or greater than one week respectively. Frequency distribution of BMRC  stage for TBM showed that out of 160 patients with TBM, 19.4% (31), 65% (104) and 15.6% 25) patients were in BMRC stage I, II and III, respectively, as presented in Figure 5 patientIn our study, 23 (14.4%) have T2DM. It was also noted in our study that 17 (10.6%) had hypertension and 26 (16.2%) were smokers.Stratification for age with respect to hyponatremia showed that 16 25%), 23 (35.9%), 06 (9.4%) and 19 (29.7%) patients who were in the age group 20-30 years, 30-40 years, 41-50 years and 51-60 years had hyponatremia, respectively. Whereas 18 (18.8%), 25 (26%), 19 (19.8%) and 34 (35.4%) patients who were in the age group 20-30 years, 31-40 years, 41-50 years and 51- 60 years did not have hyponatremia, respectively  and 62 (64.6%) who were in the male group had and did not have hyponatremia respectively. Whereas 30 (46.9%) and 34 (35.4%) who were in the female group had and did not have hyponatremia, respectively. p-value was 0.10 as presented in Table Stratification for the duration of symptoms with respect to hyponatremia showed that patients who had symptoms < 1 week, 35 54.7%) had hyponatremia. Whereas patients who had symptoms > 1 week, 29 (45.3%) had hyponatremia. p-value was 0.24 as presented in Table .7% had hStratification for BMRC stage with respect to hyponatremia showed that patients who had BMRC stage I, 10 15.6%) had hyponatremia. Whereas patients who had BMRC stage II & III, 43 (67.2%) and 11 (17.2%) had hyponatremia, respectively. p-value was 0.60 as presented in Table .6% had hStratification for diabetes mellitus type II with respect to hyponatremia showed that patients who had diabetes mellitus, 07 (10.9%) had hyponatremia. Whereas patients who did not have diabetes mellitus, 57 (89.1%) and 80 (83.3%) had and did not have hyponatremia respectively. p-value was 0.21 as presented in Table Stratification for hypertension with respect to hyponatremia showed that patients who had hypertension, 07 (10.9%) had hyponatremia. Whereas patients who did not have hypertension, 57 (89.1%) had hyponatremia. p-value was 0.55 as presented in Table Stratification for smoking status with respect to hyponatremia showed that patients who smoked, 10.7% (07) had hyponatremia. Whereas patients who did not smoke, 89.1% (57) had hyponatremia and 80.2% (77) did not had hyponatremia. p-value was 0.10 as presented in Table In this study, serum sodium levels were evaluated to determine the frequency of hyponatremia in patients presenting with TBM in order to establish the local perspective as there is paucity of local data. A total of 160 patients diagnosed with TBM were included in this study according to the inclusion criteria. Out of 160 patients with TBM, 40% (64) had hyponatremia. It was also found that 14.4% (23) had SIADH and 25.6% (41) had CSWS.A prospective hospital-based study comprising 76 patients with TBM reported that 34 (44.7%) TBM patients had hyponatremia. It was further observed that hyponatremia was due to cerebral salt wasting in 17, syndrome of inappropriate secretion of antidiuretic hormone in 3 and miscellaneous causes in 14 patients. Outcome of TBM was related to the duration of hospital stay, level of consciousness, abnormal neurological exam, mechanical ventilation, severity of infection, age and other comorbidities . KarandaCNS infections treatment and management is emerging as a great challenge especially in the setting of hyponatremia and its complication. Moreover, differentiating CSWS from SIADH is of paramount importance as both are managed paradoxically but present somewhat similarly. Data from our study would possibly offer new insight to clinicians that can impact the understanding of hyponatremia in TBM and aid in the improvement of active management strategies, improving quality of life and patient well-being.Our study had a few limitations. Our study used a smaller sample size of admitted patients compared to other studies we discussed. We also used a relatively short duration of study. Future studies with larger populations and longer duration will help mitigate these limitations.Our study showed that the prevalence of hyponatremia in patients with TBM is significant, consistent with previous studies. Therefore, as determined in our study, assessment of all patients of TBM is vital for the evaluation of hyponatremia to predict outcome and to improve management. It is also warranted as lack of appropriate evaluation and management can lead to grave and permanent neurological consequences, thereby increasing morbidity and mortality. Clinicians should be aware of the implication of sodium imbalance among patients of TBM and differentiate between the various therapeutic options in order to avoid morbidity and mortality.


Multivariate analysis was used to identify independent prognostic features for survival in LGSOC and HGSOC. Adjuvant chemotherapy was given in 99.7% of HGSOC patients versus in 27% of LGSOC (p < 0.0001). The LGSOC patients had greater surgical complexity scores (p < 0.0001), more frequent postoperative ICU/HDU admissions (p = 0.0002), and higher peri-/post-operative morbidity (p < 0.0001) compared to the HGSOC patients. The 5-year OS and progression-free survival (PFS) was 30% and 13% for HGSOC versus 57% and 21.6% for LGSOC, p = 0.016 and p = 0.044, respectively. Surgical complexity  and complete cytoreduction  were independent prognostic features for OS in LGSOC. This study demonstrates no clear significant survival advantage of chemotherapy in LGSOC. It highlights the substantial survival benefit of dynamic multi-visceral surgery to achieve complete cytoreduction as the primary treatment for LGSOC patients.In our center, adjuvant chemotherapy is routinely offered in high-grade serous ovarian cancer (HGSOC) patients but less commonly as a standard treatment in low-grade serous ovarian cancer (LGSOC) patients. This study evaluates the efficacy of this paradigm by analysing survival outcomes and by comparing the influence of different clinical and surgical characteristics between women with advanced LGSOC ( Ovarian cancer is the fourth most common cause of cancer-related death in women worldwide and is the most lethal gynecological malignancy . Since eEpithelial ovarian cancer (EOC) is categorized by diverse histological subtypes and differentiations . Serous Cytoreductive surgery, which aims at nil macroscopic residual disease (RD), is a pivotal part of treatment for ovarian cancer ,15. The This study was primarily designed to assess the differences in the clinical management strategies between patients with advanced LGSOC and those with HGSOC. The second aim of this study was to evaluate the justification of omitting standard adjuvant chemotherapy in LGSOC. By comparing the surgical characteristics and survival outcomes between low- and high-grade tumours, we hypothesised that there would be a disparity in the treatment characteristics and survival outcomes.n = 1) or recurrent ovarian malignancy (n = 5), and those who had surgery in an emergency setting (n = 3) were excluded, see Prospectively collected data from a cohort of consecutive patients who had been treated for advanced-stage serous EOC between January 2014 and December 2017 in St. James’s University Hospital, Leeds, United Kingdom, were retrieved from the hospital wide electronic patient database PPM . TreatmeFor this analysis, age was defined as age at the time of diagnosis. The clinical condition of the patients was scored by means of the performance status (PS) . All of n = 6) or who had radiologically progressed on NACT (n = 2) were counselled against adjuvant chemotherapy due to lack of benefit. A total of seven LGSOC patients received post-operative adjuvant chemotherapy only, four patients received six cycles of carboplatin and paclitaxel, and three patients received single agent carboplatin. A further three LGSOC patients received post-operative chemotherapy as part of their interval debulking regime. Other adjuvant management strategies for this LGSOC cohort included bevacizumab in combination with chemotherapy (n = 1), maintenance letrozole (n = 2), and maintenance tamoxifen (n = 2).Patients receiving NACT followed by surgical cytoreduction were those patients in which unacceptable surgical morbidity was expected, such as patients with PS > 2 or those requiring extensive preoperative optimization or those in who complete cytoreduction was unlikely to be achieved, as indicated by imaging. NACT consisted of three courses of combined carboplatin and paclitaxel chemotherapy prior to surgery with or without a subsequent three courses of adjuvant chemotherapy. A total of 13 LGSOC patients received NACT (35.1%) compared to 240 HGSOC patients (80%). Adjuvant chemotherapy in the upfront surgical cytoreduction setting usually consists of six courses of combined carboplatin and paclitaxel chemotherapy. Patients with LGSOC were offered adjuvant combined chemotherapy but were counselled regarding the chemo-resistant nature of the disease and therefore the limited lack of efficacy. LGSOC patients who had no radiological response from NACT  . The proThe primary outcome parameter of overall survival (OS) was calculated from the date of diagnosis to the date of death from the disease. The secondary parameter, progression-free survival (PFS) was calculated from the date of diagnosis to the date of confirmed recurrence. Other secondary parameters included the timing of surgical cytoreduction (PDS or IDS), PS, complexity of the surgical procedure, residual disease (RD), duration of the surgical procedure, intra-operative blood loss, utilization of the high-dependency (HDU) or intensive care unit (ICU), peri- and post-operative morbidity, and length of hospital stay. Residual disease was categorized according to the size of the remaining tumour nodules at the end of the surgical procedure. Complete cytoreduction was defined as a no measurable macroscopic RD (CC 0) or RD < 2.5 mm (CC 1). Incomplete cytoreduction was defined as CC 2 (2.5 mm ≥ RD < 2.5 cm) or CC 3 (RD ≥ 2.5 cm) . The comThe disease was staged as defined by the 2014 FIGO criteria . Data rep value < 0.1 in the univariate analysis were then combined in a multivariate analysis. All of the tests were two-sided, and p < 0.05 was considered significant. The software packages GraphPad Prism  and SPSS 27  were employed for data analysis.The characteristics of the patients according to group were presented as means +/− SD. Differences between the groups were analyzed by the Chi-squared, Student T-test, or Mann–Whitney tests, depending on the data distribution. Standard univariate analysis compared survival curves using the Kaplan–Meier method, and the statistical significance of these parameters were calculated using the log-rank test. Multivariate analyses were performed using Cox’ proportional hazard method. Independent variables that were found to have a p < 0.0001. The pre-treatment CT scan showed calcified deposits in 78.4% of the LGSOC patients compared to in 5.7% of the HGSOC patients, p < 0.0001. Details are displayed in A total of 337 patients were enrolled in the study: 37 confirmed LGSOC and 300 patients with HGSOC. There were no statistical differences in age, FIGO stage, or performance status between the two groups. The median pre-treatment CA125 measurements were 122 (range 25–9657) U/mL for the LGSOC patients versus 875  U/mL for the HGSOC patients, p < 0.0001. The median PCI in the patients with LGSOC was eight (range 2–21) versus five (range 1–24) for those with HGSOC, p = 0.0216. A complete surgical cytoreduction (CC 0–1) was achieved in 89.2% of the LGSOC patients and in 88.0% of the HGSOC patients, p = 0.833. Moderate and high SCS were found in 59.5% and 8% for the LGSOC group versus in 24.6% and 4.4% for the HGSOC group, p < 0.0001. On average, cytoreductive surgery lasted 207 ± 93 min in the LGSOC patients and 150 ± 63 min in the HGSOC patients, p = 0.0019. The average intraoperative blood loss for patients with LGSOC was 632 ± 329 cc compared to 478 ± 323 cc for those with HGSOC, p = 0.0002. An elective postoperative ICU/HDU admission was required in 37.8% of the patients with LGSOC versus in 16.3% of the patients with HGSOC, p = 0.0015. The median hospital stay length for the LGSOC patients was 9 (range 4–30) days, and it was 7 (range 7–68) days for the HGSOC patients, p = 0.0002. Median end of treatment CA125 in the patients with LGSOC was 17 (range 5–139) U/mL versus 13 (range 3–4019) U/mL in the HGSOC, p = 0.215. Clinically relevant perioperative morbidity, CD 3A or higher, was observed in 27% of the patients with LGSOC compared to in 6.6% of those with HGSOC, p < 0.0001. One patient with HGSOC died within 30 days following cytoreductive surgery due to bowel related complications. Two further patients died 60 days postoperatively, one in each group. Neither were directly related to their surgical procedure; one death was related to chemotherapy complications at 48 days, and the other died from an undisclosed accident at 45 days, respectively. The majority of patients with LGSOC (73%) did not receive adjuvant chemotherapy treatment, in contrast to only one HGSOC patient (0.3%) p < 0.0001. Details are displayed in Primary cytoreduction was performed in 64.9% of the patients with LGSOC versus in 20.0% of the patients with HGSOC, p = 0.016 (p = 0.044.The median follow-up was 70 months. The 5-year OS for the whole cohort was 33% (median OS 41 months 95% CI 35.3–46.7). The 5-year OS for patients with LGSOC was 57%  compared to 30% (median 40 months 95% CI 34.6–45.4) for those with HGSOC,  = 0.016 . The fivp = 0.001) versus 24% and 11% for the IDS patients (p = 0.001). The median OS for the LGSOC patients who underwent PDS was not reached, mean 68.6 months (95% CI 58.8–73.4) and median PFS 29 months (95% CI 6.2–51.8) versus 66 months (95% CI 48.6–83.5) and 24 months (95% CI 18.9–29) for those HGSOC patients who underwent PDS, p = 0.097 and 0.552, respectively. The median OS and PFS for the LGSOC patients who underwent IDS was 31 months (95% CI 18–44) and 16 months (95% CI 11.2–20.7) compared to 36 months (95% CI 31.6–40.4) and 16 months (95% CI 14.4–17.6) for the HGSOC group who underwent IDS, p = 0.897 and 0.874, respectively.A superior OS was established in the entire cohort with PDS when compared to those with IDS . The 5-yp < 0.0001. The median OS for the LGSOC patients with a CC of 0–1was not reached: mean OS 64.8 months (95% CI 55.3.6–74.3) and median PFS 26 months (95% CI 8.7–43.3) versus HGSOC OS 42 months (95%CI 36.3–47.7) and PFS 18 months (95% CI 16–20) for those with a CC of 0–1, p = 0.012 and 0.016, respectively  was achieved compared to in those with an incomplete cytoreduction (CC ≥ 2). The 5-year OS was 32.8% for the patients with a CC of 0–1versus 0% for the patients with a CC ≥ 2, ectively . The medectively .p < 0.002. The median OS for the LGSOC patients with an SCS ≥ 4 was not reached, with mean an OS of 66.5 (95% CI 55.3–77.7) and a median PFS of 36 (95% CI 25–47) months versus 50 (95%CI 37.7–62.3) and 21 (95%CI 15–26) months for those HGSOC patients with an SCS ≥ 4, p = 0.019 and 0.132, respectively  when compared to those who had low surgical complexity (SCS 1–3). The 5-year OS was 44 % for patients with an SCS ≥ 4 versus 24 % for patients with an SCS 1–3, ectively . The medectively .p < 0.1) were PS, FIGO stage, PDS, CC 0–1, SCS ≥ 4, and CD < 3. The favorable prognostic features for the HGSOC patients were the same, with the exception of FIGO stage. In the multivariate analysis, the advantageous prognostic features for OS in the LGSOC patients were a CC of 0–1  and an SCS ≥ 4 . For the HGSOC patients, the favorable prognostic features that were determined in the multivariate analysis were PDS  and a CC of 0–1 . Further details are displayed in For the LGSOC patients, the favorable prognostic features for OS that were determined in the univariate analysis  that is not commonly used to describe ovarian cancer debulking surgery but that is becoming standard practice in surgical oncology. It is classified by more discrete variables that aim to improve the precision of residual disease status reporting and has been used in other ovarian cancer survival analysis studies . The satPatients with LGSOC had a lower pre-treatment CA125 level and more frequent calcified deposits on the CT scan when compared to the patients with HGSOC. This observation has been demonstrated in other studies ,37. RemaIn addition to the aforementioned differences between LGSOC and HGSOC, there is also a clear disparity in surgical management. The finding that the LGSOC patients in our study required more complex surgical procedures (aiming at CC 0–1) when compared to those with HGSOC has, to our knowledge, not been reported before. The ramifications of more complex surgery in LGSOC were longer procedural time, more intra-operative blood loss, more HDU/ICU admissions, longer hospital stay, and consequentially, increased peri-/post-operative morbidity. This has been validated by a previous study that demonstrated a correlation between a higher PCI score and prolonged surgical time, increased post-op complications, and extended hospitalization . AdditioThe findings of our study should not be overstated. Confounding variables are the relatively small number of patients with LGSOC who were recruited over a timeframe of 4 years and the non-randomized character of the study. Furthermore, two patients did demonstrate a radiological response to solid disease while on NACT, although this was restricted to a modest reduction in omental cake, thus supporting chemoresistance, and it does not confirm total futility in all patients. Equally, data on patients with LGSOC are not widely available, making randomization almost unachievable. We also acknowledge the fact that our 20% rate of primary cytoreductive surgery for advanced EOC patients falls short of our predetermined ambition.Our study supports the consideration of omitting adjuvant systemic chemotherapy treatment for the management of advanced-stage LGSOC. The survival benefit of LGSOC compared to HGSOC is maintained despite the lack of chemotherapy, highlighting the significance of surgical treatment for this disease entity. Radical multi-visceral surgery that aims at complete surgical cytoreduction should be considered to be the primary treatment for LGSOC patients. A clear positive association between increased surgical complexity and superior survival outcomes in LGSOS was demonstrated. Nevertheless, increased operation time, longer hospital stays, increased HDU/ICU requirements, and increased post-operative morbidity resulting from this paradigm of surgical management should also be taken into consideration. LGSOC and HGSOC are different entities of serous EOC and consequently deserve different management. Therefore, we strongly recommend stratifying future studies in EOC to these separate subtypes. Prospective clinical trials should maintain an emphasis on systemic therapies that target specific LGSOC pathways such as MEK or PI3K inhibitors. Additional translational research is required to identify molecular predictors of response for LGSOC-targeted systemic treatment and endocrine therapy.