RNA interference (RNAi) is a useful tool to assess gene function by knocking down expression of a target gene and has been used successfully in domestic and laboratory organisms. However, the use of ...RNAi for functional genomics has not fully extended into ecological model organisms in natural environments. Assessment of gene function in the wild is important because gene function can be environmentally and context dependent. Here, we present a case study using RNAi to assess gene function in wild paper wasps Polistes metricus, to test roles for two candidate genes (NADH dehydrogenase (NADHdh) and retinoid and fatty acid binding protein (RfaBp)) in the development of reproductive castes. Previous studies have shown that these genes are upregulated in larvae that become queens compared to workers, but this pattern was reversed in the laboratory, making field-based studies necessary. We orally administered dsRNA to larvae in field colonies and found evidence of a short-term knockdown followed by a compensatory rebound in expression for RfaBp. We also observed the predicted worker-like decrease in lipid stores in NADHdh dsRNA treated wasps, suggesting a possible role for NADHdh in caste development. We discuss our results in the context of challenges for using RNAi for functional genomics in ecological model organisms in the field.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
In contrast to endometritis, now diagnosed by cytological examination, the effect of endocervical inflammation on reproductive performance has been inadequately investigated. In this study, ...endocervical and endometrial cytological specimens were collected from 168 Holstein cows between 21 and 60 days in milk (DIM) to investigate the prevalence of endocervical inflammation and effect on days to conception. Statistical analyses were stratified based on DIM at examination (<35 vs. ≥35 DIM). Endocervical inflammation with ≥5% neutrophils before 35 DIM (disregarding the level of endometrial inflammation) was associated with decreased hazard of pregnancy within 300 DIM (adjusted hazard ratio, 0.4; 95% confidence interval: 0.3–0.8). A decrease in hazard of pregnancy was observed when >6% neutrophils were counted in endometrial smears (adjusted hazard ratio, 0.4; 95% confidence interval: 0.2–0.7). The study suggested an additive effect of combined endocervical and endometrial inflammation on the hazard of pregnancy within 300 DIM. Using the thresholds of 5% neutrophils for the cervix and 6% neutrophils for the uterus, 11% of the cows examined before 35 DIM presented cervicitis only, 13% were affected by endometritis only, and 32% suffered from both endometrial and endocervical inflammation. The presence (absence) of cervicitis was not indicative of the presence (absence) of endometritis. This study showed that in addition to uterine inflammation, endocervical inflammation in early lactation affects conception. Thus, the global evaluation of genital tract health may be more beneficial for reproductive performance than that of endometrial inflammation.
Cattle besnoitiosis caused by Besnoitia besnoiti (Eucoccidiorida: Sarcocystidae) is a re‐emerging disease in Europe. Its mechanical transmission by biting flies has not been investigated since the ...1960s. The aim of this study was to re‐examine the ability of Stomoxys calcitrans (Diptera: Muscidae) to transmit virulent B. besnoiti bradyzoites from chronically infected cows to susceptible rabbits. Three batches of 300 stable flies were allowed to take an interrupted bloodmeal on chronically infected cows, followed by an immediate bloodmeal on three rabbits (Group B). A control group of rabbits and a group exposed to the bites of non‐infected S. calcitrans were included in the study. Blood quantitative polymerase chain reaction (qPCR) analyses, and clinical, serological and haematological surveys were performed in the three groups over 152 days until the rabbits were killed. Quantitative PCR analyses and histological examinations were performed in 24 tissue samples per rabbit. Only one rabbit in Group B exhibited clinical signs of the acute phase of besnoitiosis (hyperthermia, weight loss, regenerative anaemia and transient positive qPCR in blood) and was seroconverted. Parasite DNA was detected in four tissue samples from this rabbit, but no cysts were observed on histological examination. These findings indicate that S. calcitrans may act as a mechanical vector of B. besnoiti more efficiently than was previously considered.
This is the first reproduction of the mechanical transmission of Besnoitia besnoiti by stable flies in a heterologous model.
A batch of 300 stable flies was sufficient to transmit virulent B. besnoiti bradyzoites from chronically infected cattle to rabbit.
Transmission was effective: one infected rabbit exhibited clinical signs of acute besnoitiosis and seroconversion.
Background
Reference intervals are the most common tool used to interpret results of laboratory tests. However, in veterinary clinical pathology, the number of available reference individuals is ...often small.
Objectives
The purpose of this study was to investigate the effects of small reference sample groups on the imprecision of the reference limits.
Methods
Gaussian and log‐Gaussian distributions of 10 ≤ n ≤ 750 values were analyzed. Reference limits and 90% confidence interval of limits (90% CI) were calculated. Imprecision of limits was estimated by the ratio of the width of the 90% CI: width of the reference interval (WCI/WRI).
Results
For Gaussian distributions, the WCI/WRI ratio cannot be expected to be lower than 0.2 when n < 55. In log‐Gaussian distributions, the ratio greatly increases for the upper limit with skewness toward high values, whereas it moderately decreases for the lower limit.
Conclusion
Independent of the size of the reference sample group, it is very important to report the CIs of the reference limits, which can be very large for small reference sample groups. When the sample size is very small (n < 20), calculations maybe misleading and it is better to instead report all values.
Background: According to international recommendations, reference intervals should be determined from at least 120 reference individuals, which often are impossible to achieve in veterinary clinical ...pathology, especially for wild animals. When only a small number of reference subjects is available, the possible bias cannot be known and the normality of the distribution cannot be evaluated. A comparison of reference intervals estimated by different methods could be helpful.
Objective: The purpose of this study was to compare reference limits determined from a large set of canine plasma creatinine reference values, and large subsets of this data, with estimates obtained from small samples selected randomly.
Methods: Twenty sets each of 120 and 27 samples were randomly selected from a set of 1439 plasma creatinine results obtained from healthy dogs in another study. Reference intervals for the whole sample and for the large samples were determined by a nonparametric method. The estimated reference limits for the small samples were minimum and maximum, mean ± 2 SD of native and Box–Cox‐transformed values, 2.5th and 97.5th percentiles by a robust method on native and Box–Cox‐transformed values, and estimates from diagrams of cumulative distribution functions.
Results: The whole sample had a heavily skewed distribution, which approached Gaussian after Box–Cox transformation. The reference limits estimated from small samples were highly variable. The closest estimates to the 1439‐result reference interval for 27‐result subsamples were obtained by both parametric and robust methods after Box–Cox transformation but were grossly erroneous in some cases.
Conclusion: For small samples, it is recommended that all values be reported graphically in a dot plot or histogram and that estimates of the reference limits be compared using different methods.
Background
Breed‐specific reference intervals are of increasing interest in veterinary medicine. The health monitoring of the Dogue de Bordeaux, a breed predisposed to familial juvenile ...glomerulonephropathy and hypothyroidism, would benefit from specific reference intervals.
Objective
The purpose of this study was to establish breed‐specific biochemical reference intervals for the Dogue de Bordeaux in accordance with the International Federation of Clinical Chemistry and Clinical and Laboratory Standards Institute guidelines.
Methods
One hundred and twenty Dogues de Bordeaux from France and Belgium were recruited. Complete urinalysis and chemistry panels, venous blood gas variables, total thyroxin and thyroid stimulating hormone, and fibrinogen and antithrombin were measured for each dog. Reference intervals were determined using the non‐parametric method. Confounding variables such as sex, age and color of facial mask were analyzed.
Results
Due to pre‐defined criteria for exclusion, 62 healthy dogs were finally selected for the reference intervals determination. Using the instrument manufacturer's generic canine RI for most analytes did not have a significant impact on potential clinical decisions, except for total proteins, ALT, AST, total cholesterol, lipase and total thyroxin, for which possible clinically relevant differences were noted.
Conclusion
Specific reference intervals for biochemical analytes in the Dogue de Bordeaux were determined under controlled pre‐analytical and analytical conditions, and according to international recommendations. The use of these breed‐specific reference intervals is recommended when using the specified analytic instruments, especially for the 6 analytes for which the reference intervals differed considerably from those provided by manufacturers.
Background
There is an increasing interest for breed‐specific reference intervals in veterinary medicine. In a previous study, breed‐specific biochemical reference intervals (RIs) have been ...established for Dogues de Bordeaux (DDBs). This breed is predisposed to familial juvenile glomerulonephropathy and hypothyroidism, and would benefit from hematologic RI.
Objective
The purpose was de novo establishment of breed‐specific hematologic RIs for the DDB in accordance with the International Federation of Clinical Chemistry and Clinical and Laboratory Standards Institute guidelines.
Methods
One hundred and twenty DDBs from France and Belgium were recruited. CBCs were determined with the Sysmex XT‐2000iV analyzer within 12 hours of blood collection. RIs were determined using the nonparametric method. Effects of sex, age, and face mask color were studied.
Results
RIs were determined in 58 healthy dogs. DDBs had higher RIs for HGB, HCT, MCV, MCHC, and mean platelet volume, and lower RIs for reticulocytes counts, platelets by impedance (PLT‐I) and optical count (PLT‐O), and plateletcrit when compared with generic canine RIs. Age significantly affected RIs for HGB, HCT, MCHC, WBC, neutrophil, lymphocyte, and monocyte counts.
Conclusion
The generic canine RIs established in the same laboratory with analogous preanalytical and analytical variations did not differ significantly from breed‐specific RIs, and thus have no significant impact on clinical decision making; however, breed‐specific RIs are advised for some RBC and all platelet‐related variables to avoid erroneous suspicion of polycythemia and thrombocytopenia when using general canine RIs for evaluation of DDB.
Background
Reference intervals are important aids for interpreting clinical pathology laboratory data especially in Cynomolgus monkey (Macaca fascicularis), the non‐human primate species most widely ...used in biomedical research. The purpose of this study was to establish hematologic reference intervals for Cynomolgus according to the International Federation of Clinical Chemistry and Clinical and Laboratory Standards Institute guidelines using the databank at a primatology center.
Methods
Blood specimens from 272 healthy Cynomolgus imported from Mauritius, the Philippines and Vietnam, were analyzed. Reference intervals were established by nonparametric method. Effects of sex, age, body weight, and breeding origin were investigated.
Results
Hemoglobin, mean corpuscular hemoglobin, and mean corpuscular hemoglobin concentration decreased slightly and mean corpuscular volume increased slightly with age. Lower red blood cell concentration, hemoglobin, and hematocrit were observed in monkeys from the Philippines.
Conclusions
These hematology reference intervals, established according to international recommendations, can be used in settings using similar animals and analyzers.
Reference limits are some of the most widely used tools in the medical decision process. Their determination is long, difficult, and expensive, mainly because of the need to select sufficient numbers ...of reference individuals according to well-defined criteria. Data from hospitalized patients are, in contrast, numerous and easily available. Even if all the information required for a direct reference interval computation is usually not available, these data contain information that can be exploited to derive at least rough reference intervals.
In this article, we propose a method for the indirect estimation of reference intervals. It relies on a statistical method which has become a gold-standard in other sciences to separate components of mixtures. It relies on some distributional assumptions that can be checked graphically. For the determination of reference intervals, this new method is intended to separate the healthy and diseased distributions of the measured analyte. We assessed its performance by using simulated data drawn from known distributions and two previously published datasets (from human and veterinary clinical chemistry).
The comparison of results obtained by the new method with the theoretical data of the simulation and determination of the reference interval for the datasets was good, thus supporting the application of this method for a rough estimation of reference intervals when the recommended procedure cannot be used.
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