In vivo and in vitro rearing of Pediculus humanus capitis (Anoplura: Pediculidae)

Four geographically distinct colonies of the human head louse, Pediculus humanus capitis De Geer (Anoplura: Pediculidae) were reared on a live host and exhibited significantly different life history patterns. Florida head lice exhibited approximately 10% slower development and approximately 15% reduced longevity relative to California or Ecuador head lice. Fecundity (4.9 +/- 0.2 eggs/female/d) and fertility (76.4 +/- 2.9% mean hatching rate) declined over the lifetime of female lice, especially when separated from males (i.e., unmated recently). All four colonies (above plus one from Panama) were similar in their ability to tolerate starvation, although older stages tended to die sooner. An in vitro feeding apparatus was developed to rear head lice. Teneral first instar lice were placed on human hair tufts on the upper side of membrane-covered feeders, which were immersed bottom-side down within a vessel containing warmed human blood. Relative to lice reared on a human host, in vitro-reared lice required a significantly longer time (10-20%) to molt and survived a significantly shorter time as adults (30-50%); the addition of antibiotics did not adversely affect louse development. Teneral first instars were more likely than any other stage to feed through the membrane. Lice spent a significantly greater proportion of time searching in the in vitro apparatus than on a host, but the proportion of time spent feeding did not differ. This research is the first to demonstrate that head lice can be reared successfully in vitro through a complete life cycle.     

Prevalence of Streptococcus pneumoniae in Respiratory Samples from Patients with Tracheostomy in a Long-Term-Care Facility

We aimed to study the prevalence of Streptococcus pneumoniae in respiratory samples from institutionalized patients with chronic tracheostomy. A total of 264 pairs of nasopharyngeal and endotracheal cultures were collected. There was no difference in the proportion of positive cultures between children (21%) and adults (18%). However, the proportion of positive endotracheal cultures was higher than that of nasopharyngeal cultures in adults (18% versus 3%, respectively; P < 0.001) but not in children (17% in both sites).     

Multiplex interphase FISH as a screen for common aneuploidies in spontaneous abortions

BACKGROUND: A multiplex fluorescence in-situ hybridization (FISH) strategy using chromosome-specific probes for eight chromosomes as an initial screen for chromosome abnormalities in uncultured tissues from spontaneous abortions was evaluated. METHODS: Fifty-seven prefetal spontaneous abortions were studied by karyotyping cultured cells and using FISH on uncultured cells. Two probe sets were used, identifying chromosomes 13, 15, 16, 18, 21, 22, X and Y. RESULTS: Abnormalities were detected in 53% of cases by karyotyping, and 54% of cases by FISH. FISH detected an abnormality in four of five cases where cultures failed, and in two cases where maternal cells apparently overgrew the culture. FISH missed four trisomies not identifiable with the probe sets, and one trisomy because one probe set was unscorable. FISH using these probes identified 83% of all abnormalities detected by karyotyping. CONCLUSIONS: FISH can detect abnormalities in a significant proportion of cases where the culture fails to grow or is contaminated by maternal cell growth. Multiplex FISH as an initial screen, followed by culture and karyotyping in cases where no abnormality is detected, would identify a higher proportion of chromosome abnormalities in spontaneous abortion specimens than karyotype analysis alone.     

Efficiency of sex pre-selection of spermatozoa by albumin separation method evaluated by double-labelled fluorescence in-situ hybridization

To evaluate the separation efficiency of Ericsson's two- and three- layer albumin separation methods, semen samples from 21 healthy males were studied. Seven patients already had two or more sons, another seven had two or more daughters and the other seven had primary infertility due to female factors. The semen samples were divided into three aliquots: one remained unprocessed initially, the other two aliquots went through two- and three-layer albumin separation methods respectively. All samples were then stained with X-Y double staining probes. In each group, four or five samples were processed at room temperature, and two or three at body temperature (37 degrees C). The labelling efficiency of X-Y double staining probe was over 99%. The X:Y sperm ratios were even in the original samples. The ratios of the X and Y spermatozoa were altered slightly but significantly after the two- layer (P < or = 0.05) or the three-layer (P < or = 0.005) separation. The alterations occurred only at room temperature. The X spermatozoa increased and the Y spermatozoa decreased, both to a small degree of difference (1.4-3.5%). Double fluorescence in-situ hybridization analysis therefore showed that albumin separation methods do not enrich Y spermatozoa.      

Asthma management and perceptions in rural South Africa.

BACKGROUND: Many First World countries have endeavored to measure the impact of asthma on individuals with asthma and, in addition to this quality of life evaluation, have attempted to define the quality of care for this common chronic illness. OBJECTIVE: The primary objective of this research probe was to assist the National Asthma Program in South Africa with the formulation and delivery of its outreach program to rural asthmatic patients. METHODS: A discussion/questionnaire document was compiled by Partners in Research from established literature. All interviews were conducted in either the clinics, hospitals, or respondents' homes. Both adult asthmatic patients and parents of pediatric asthmatic patients were interviewed. Interviewing took place at seven rural health clinics across South Africa. Each interview included extensive demographic details, questions on asthma definition, symptoms and symptom triggers, family history, age at diagnosis, frequency of symptoms, and treatment. RESULTS: Thirty-five adult asthmatic patients and 27 parents of pediatric asthmatic patients were interviewed. Of the adults, 40% reported wheezing at least once a week (despite diagnosis and treatment) and 19% of children reported similar symptom exacerbations. Fifty-one percent of adults and 56% of children were awakened at least once a week by cough or wheeze. Quality of life measurement reflected that, on average, 37% of responders were frightened during an acute asthma attack, and 68% of parents reported fearing the death of their asthmatic children. Fifty-one percent of adults and 33% of children had been hospitalized at least once for asthma. Although respondents claimed regular training in use of inhaler device, only 43% of adults completed each step correctly. CONCLUSIONS: There is a great deal of fear and ignorance surrounding asthma and, therefore, there is a real need for a greater level of patient education even in the rural areas of South Africa. In rural South Africa, attention should be paid to nurses, because they play a greater role than doctors in management and education of asthma.     

Effect of delayed specimen processing on cytomegalovirus antigenemia test results.

Blood samples held at either 4 degrees C or room temperature for 1 day had similar mean decreases in number of cytomegalovirus antigenemia-positive cells (52 to 55%) and similar false-negative test results (13 to 14%). After 2 days, samples held at 4 degrees C showed no further decline, whereas samples held at room temperature had a mean 81% decrease in positive cells, a 32% false-negative rate, and a more marked deterioration in cell morphology.     

Effect of sublethal dose of diflubenzuron and azadirachtin on various life table attributes of Culex quinquefasciatus (Diptera: Culicidae)

The exposure of sublethal doses (25% emergence inhibition) of diflubenzuron (DFB) (0.0002 mg/liter) and azadirachtin (AZD; 0.0079 mg/liter) insect growth regulators was provided to late third- early fourth instars of Culex quinquefasciatus till emergence of adults to find out the effects on various biological and life table attributes of adults, and comparison was made with control (CTL) as well. Life expectancy of both males and females of DFB survivors was significantly less in comparison with AZD and CTL ones; however, it was nonsignificantly different between CTL and AZD. In all treatments, males were shorter lived than females of their respective treatments. DFB survivors showed significant reduction in number of blood-fed females, resulting in minimum of oviposition days, total number of egg rafts, total number of eggs, eggs per egg raft, eggs per female life span, and eggs per female per day than survivors of both AZD and CTL significantly, but these parameters did not differ significantly between CTL and AZD, except eggs per egg raft. The net reproductive rate, intrinsic rate of increase, and finite rate of increase were significantly less in DFB than rest of the survivors, whereas doubling time for generation was significantly high; however, difference in mean generation was not significant among them. This study reveals that DFB has the potency to affect severely the various life table attributes at sublethal dose, but AZD does not have such effectiveness at similar sublethal dose against Cx. quinquefasciatus.     

The effects of early rearing environments on maternal behavior in adult female rats

The purpose of this study was to determine the effect of presence of siblings and exposure to colony cues during early life on the onset of maternal behavior at 20 and 75 days of life, and on subsequent emotional behavior in juvenile and adult female rats. Different groups of animals were reared either with a large number or a small number of siblings (sibling factor), or in the colony room or in a room apart from other mothers with pups (colony factor). Animals were tested for latency (in days) to show maternal behavior at either 20 or 75 days of age, and for emotional behavior (using the open field apparatus) at either 20-40 or 80-100 days of age. Exposure to colony sensory cues significantly reduced latencies for pup retrieval and nest-building in adults, but not in juveniles. Exposure to a large number of siblings reduced the latencies, in both adults and juveniles, to nest build; in juveniles, to lie over 75% of the litter; and in adults, to adopt a nursing posture. Consistent with prior research, juveniles had longer latencies than adults for a number of pup-directed behaviors including: latency to become maternal, latency to retrieve pups, latency to adopt a lactating posture, latency to lick/genital lick, and latency to nest-build. In the emotionality tests, the young animals scored as less emotional than the adults, an effect which was further exaggerated by early colony exposure. Overall results indicated that manipulation of early rearing environments does have a significant positive effect on later responses to young pups by both juvenile and adult female rats.     

Anaplasmosis in Uganda. I. Use of Dried Blood on Filter Paper and Serum Samples for Serodiagnosis of Anaplasmosis—a Comparative Study

The suitability of blood collected on filter papers in comparison with corresponding conventional serum samples in the diagnosis of bovine anaplasmosis was studied using the Complement Fixation Test (CFT), DOT-ELISA, Western immunoblot and Rapid Card Agglutination Test (RCAT). Dried blood on Whatman filter paper no. I was eluted in 1.8 ml of PBS 0.05% Tween 20 given an initial dilution of 1:100.
The reactivity in both DOT-ELISA and Western immunoblotting was similar to that obtained with the sera diluted 1:100. Filter paper samples gave lower reactivity in all the tests as compared with corresponding serum samples. There was no significant difference in the reactivity between the eluates from filter papers stored at room temperature and those stored at 4°C. Storage at room temperature did not significantly affect reactivity for up to 6 months. Eluates from filter papers stored for 6 months at room temperature continued to give similar reactivity to those from freshly prepared filter papers in both DOT-ELISA and Western blot, and in the Rapid Card Agglutination Test.
It is concluded that collecting blood on filter papers is a suitable technique for large-scale screening and for seroepidemiological studies on anaplasmosis, and offers many advantages especially in developing countries where transport and cold chain facilities are a major constraint.     

An automated feeding apparatus for in vitro maintenance of the human head louse, Pediculus capitis (Anoplura: Pediculidae)

An automated feeding apparatus was developed to maintain the human head louse (Pediculus capitis DeGeer) in vitro. With the use of valves and timers, banked human blood and saline from refrigerated reservoirs were pumped into and flushed out of the system every 7 d. During this rotational interval, bloodmeals were provided to head lice continuously and ad libitum through a stretched Nescofilm-silicone sandwich membrane. Compared with our previous in vitro human head louse-rearing apparatus, greater numbers of lice could be fed simultaneously with minimal human monitoring. Development of second to third instars and third instars to adults was significantly faster when lice were reared in vivo than on either of the in vitro rearing systems; there was no significant difference in the duration of the first instar. Although fecundity and hatch rates were significantly higher for female lice reared in vivo, similar trends have been observed for other membrane-fed arthropods. Body lice (Pediculus humanus L.) and bed bugs (Cimex lectularius [L.]) also completed most of their life cycle on this apparatus. Our automated mass-rearing system has broad applications for maintaining fluid-sucking ectoparasites and will facilitate various toxicological, behavioral, and disease-transmission investigations.     

Pre-test blood samples at clinical laboratories--effects of temperature and duration of blood sample storage on measurements

Increases in lactate dehydrogenase (LD) were observed from 24 hours for samples stored at normal temperature, and were noted from 12 hours for those stored at room temperature. Potassium (K) increased from 3 hours for samples stored under refrigeration, and increased from 6 hours for those stored at normal temperature. For those stored at room temperature, this parameter decreased from 6 through 12 hours after blood sampling and increased from 24 hours. Inorganic phosphorus (P) increased from 36 hours for samples stored at normal temperature. For those stored at room temperature, this parameter decreased from 3 through 6 hours after blood sampling and increased from 24 hours. Increases in magnesium (Mg) were noted from 24 hours for samples stored at normal temperature, and were observed from 12 hours for those stored at room temperature. Increases in iron (Fe) were noted from 36 hours for samples stored at normal temperature, and were observed from 24 hours for those stored at room temperature. Blood urea nitrogen(BUN) increased from 24 hours for samples stored at room temperature. Creatinine (CRE) increased from 24 hours for samples stored at room temperature. Uric acid (UA) decreased from 36 hours for samples stored at room temperature. Serum glucose decreased from 6 hours for samples stored under refrigeration, decreased from 6 hours for those stored at normal temperature, and decreased from 3 hours for those stored at room temperature. Although it has generally been accepted that it may be good practice to store blood samples under refrigeration, there have been no established theories to prove this. The authors investigated the effects of the temperature and duration of blood sample storage on measurements and found that for some parameters, the measurements varied depending on the temperature and, duration of storage. No specific temperature for the storage of blood samples could be identified in all cases and accordingly, the personnel concerned should refer to the relevant materials and documents so as to identify an appropriate storage temperature.     

Receptors for immunoglobulin and complement on sheep RBC-binding lymphocytes in newborn infants.

Sheep erythrocyte-binding human peripheral blood lymphocytes bearing receptors for the Fc-part of IgG and/or complement were studied in twenty newborn pre-term and term infants and fifteen adult controls. A technique with mixed rosettes and fluorescein-labelled sheep erythrocytes was used. The newborn infants had a significantly lower proportion of lymphocytes with receptors for sheep erythrocytes and the Fc-part of IgG (1.6%) than the adults (6.7%). Also the percentage of lymphocytes binding both sheep erythrocytes and complement-coated indicator cells was significantly lower in the newborn infants (2.6%) than in the adults (8.9%). A significantly higher proportion of lymphocytes had receptors for sheep erythrocytes and complement than for sheep erythrocytes and the Fc-part of IgG in both newborn infants and adults. There was no correlation between the values of lymphocytes binding sheep erythrocytes, or the Fc of IgG or complement and the proportion of mixed rosettes.     

Effect of delay in processing and storage temperature on diagnosis of SARS-CoV-2 by RTPCR testing

PurposeA large number of new molecular or virology laboratories have been established to increase the testing capacity for SARS-CoV-2. Due to heavy workload, there is delay in testing of samples. In order to avoid the negative effect of delayed testing on RTPCR results guidelines are issued from WHO and CDC to transport samples in cold chain. However, in pandemic situations the recommended guidelines for transport and storage conditions are often compromised. This study was conducted to evaluate the effect of sample storage conditions at different temperatures on the results of RT PCR test.MethodsTotal 275 samples were included in this study, among these 126 samples tested positive and 149 samples tested negative. All samples were aliquoted into two and the aliquotes stored in duplicate at 4 ?°C and room temperature. All aliquots stored at both the temperatures were tested by RTPCR every 24 hours up to 5 days.ResultsDiagnostic accuracy decreased from day1 to day 5 at both the storage temperatures i,e 4 ?°C and room temperature in comparison to the initial day results. Positivity decreased on an average of 9.02% at 4 ?°C and at 9.27% at room temperature per day. Among total 126 positive samples on an average false negative and failure of internal control at 4 ?°C and room temperature was 8.86%, 8.22% and 3.64%, and 4.12%, respectively. All the samples with CT value ?< ?30 remained positive at both temperatures up to 5 days. Few samples with >30 CT value showed variable results i.e. positive, negative or internal control failure from day 1 (2nd day after sample collection) onwards.ConclusionsThere was no significant difference between RT PCR results of samples stored at 4 ?°C and room temperature up to 5 days of collection. However internal control failure was more in samples stored at room temperature. Therefore, samples received without cold chain also may be processed by RTPCR and should not be rejected.     

Temperature-dependent development and survival rates of Culex quinquefasciatus and Aedes aegypti (Diptera: Culicidae)

Development, growth, and survival of Culex quinquefasciatus Say and Aedes aegypti (L.) were determined at six constant temperatures (15, 20, 25, 27, 30, 34 degrees C). The Sharpe & DeMichele four-parameter model with high-temperature inhibition described the temperature-dependent median developmental rates of both mosquito species. In both species, body size generally decreased as temperature increased. Head capsule widths in all instars in both species were significantly greater at 15 than at 30-34 degrees C. Except for the third instar of Ae. aegypti, the larval body lengths in both species were significantly greater at 15 than at 34 degrees C. All instars and pupae of both species and the adults in Cx. quinquefasciatus were significantly heavier at 15 than at 27-34 degrees C. In Cx. quinquefasciatus, survival from eclosion to adult emergence was highest in the range from 20 to 30 degrees C (85-90%) and dropped drastically at 15 (38%) and 34 degrees C (42%). In Ae. aegypti, survival to adult stage was high at 20 (92%) and 27 degrees C (90%) and lowest at 15 degrees C (3%).     

Optimization of the fluorescence in situ hybridization (FISH) technique for high detection efficiency of very small proportions of target interphase nuclei

Using commercially available fluorochrome-labeled probes specific for chromosomes X, Y, 13, 18, and 21, we optimized the technical protocols for fluorescence in situ hybridization (FISH) so that the highest sensitivity and specificity were achieved. Also, we compared the optical properties of different types of fluorescent labels in an effort to develop the most efficient FISH protocol, including the determination of which types of labels are the easiest to count accurately. The lymphocytes were purified from blood of normal male and female newborns, normal male and female adults, and a trisomy 21 male adult. Male and female lymphocytes were mixed in five different combinations. For each combination, the male lymphocytes either from newborns or from adults were diluted with female lymphocytes in seven different proportions. For each of these 35 different cell mixtures, 100,000 nuclei were analyzed and scored in a blind fashion. Among the different fluorochrome-labeled probes, the highest sensitivity and specificity were achieved when SpectrumAqua CEP-Y/SpectrumOrange CEP X probe mixture, SpectrumAqua CEP-18, SpectrumOrange LSI-13, and SpectrumOrange LSI-21 were hybridized. The hybridization sensitivity and specificity were higher than 99% for the identification of chromosomes X, Y, 13, and 18, and higher than 98% for the detection of trisomy 21. The proportion of false-positive signals was under 0.005% for XY detection and lower than 0.14% for autosome detection. With these high hybridization sensitivities and specificities, the optimized FISH protocol developed in our laboratory has the potential to detect very rare events, e.g., when the proportion of cells being sought is lower than 0.01%. In other words, our protocol allows the specific detection of one male cell sunken among 10,000 female cells.     

Long-term stability of frontal electroencephalographic asymmetry in adults with a history of depression and controls.

We investigated the stability in resting EEG across a 1- to 3-year interval in 49 adults (33 female and 16 male) with a history of unipolar depression (first onset prior to the age of 14) and 50 controls (33 female and 17 male) with no history of major psychopathology. Current depressive symptoms were quantified by self-report at both assessments. For the entire sample, EEG asymmetry in the alpha range was moderately stable (intraclass correlations between 0.39 and 0.61). Sex, history of depression, depressive symptom severity at Time 2, and change in symptom severity between Time 1 and Time 2 were unrelated to stability of EEG asymmetry. These findings support the view that resting frontal EEG asymmetry reflects a moderately stable individual difference in adults, irrespective of sex and history of depression.     

Population ecology and movement of the American cockroach (Dictyoptera: Blattidae) in sewers

The population size, age-class structure, and movement of the American cockroach, Periplaneta americana (L.) (Dictyoptera: Blattidae), were studied in three sewers in Penang, Malaysia, from September 2008 to October 2009. Eighteen to 20 glass-jar traps (two per manhole) were deployed for a 24-h period during each sampling occasion at each sewer. Adults and nymphs were active throughout the study period, with an average monthly trap catch of 57-97 adults and 79-99 nymphs. The mean proportions of adults and nymphs at the three sewers ranged from 0.47 to 0.57. Of the 2177 male and 2717 female cockroaches marked and released over the three sewers, recapture rates were 29.4-45.8 and 30.8-47.0%, respectively. The proportion of marked males and females did not differ significantly from the proportion of recaptured marked males and females. However, the mean number of times a marked female was recaptured was significantly greater than that of males. Of the 783 males and 1,030 females that were marked and recaptured, 19.4 and 24.7%, respectively, had moved between manholes, and significantly more females than males moved between manholes. Of the 406 recaptured marked adults that moved between manholes, 90.4% moved a distance of 2-20 m from their initial release site; one male moved 192 m, the longest distance recorded. Trap catch on each sampling occasion was positively correlated with daily mean temperature. The number of cockroach movements between manholes also was correlated with the mean daily minimum temperature.     

印鼠客蚤实验室种群的羽化规律研究

为开展蚤传疾病的传播机制、蚤的生理毒理、杀蚤剂的药物开发等研究提供材料．我们用印鼠客蚤的实验室种群进行蚤的羽化规律研究。该种群用人工饲料饲养幼虫,小白鼠作为供血动物。结果表明,在每1000只蚤的繁殖体系中,供血10天后,子代蚤羽化开始的前4天,雌虫占全部成虫的98．6％;羽化的第9、10天为高峰,雌性的日羽化指数（日羽化率×日雌性比）分别为7．3％和6．7％,第14天以后逐渐下降;羽化开始的第5天后,雄蚤的羽化量逐渐增加,到第14天达到高峰．日羽化指数占全部成蚤的5．9％。成蚤日累计羽化率随时间的变化为“S”型。雌性印鼠客蚤的羽化数量与时间呈负相关。可见,印鼠客蚤的雌性个体比雄性发育快,分别存在不同的羽化高峰,两个高峰重叠,延长成蚤羽化高峰日。关键词印鼠客蚤;实验室种群;羽化规律;性比     

Air bubbles and temperature effect on blood gas analysis.

The effect of temperature, time of storage, and presence of air bubbles in specimens for blood gas analysis was studied. The results show that air bubbles in a 10% proportion are undesirable because of significant elevation in the PO2, and the storage of anaerobic blood samples at room temperature (25 degrees C) is acceptable when measurements are done within the first 20 minutes.