Numbers Of Cryptosporidium Parvum Oocysts Research Articles

Polyphenolic Profile of Herniaria hemistemon Aerial Parts Extract and Assessment of Its Anti-Cryptosporidiosis in a Murine Model: In Silico Supported In Vivo Study.

Herniaria hemistemon J.Gay is widely used in folk medicine to treat hernia. The present study aimed to annotate the phytoconstituents of H. hemistemon aerial-part extract and investigate its in vivo anticryptosporidial activity. The chemical characterization was achieved via the LC-ESI-MS/MS technique resulting in the annotation of 37 phytocompounds comprising flavonoids and phenolic acids. Regarding the anticryptosporidial activity, fifty dexamethasone-immunosuppressed mice were separated into five groups: GI, un-infected (normal control); GII, infected but not treated (model); GIII, infected and received NTZ, the reference drug; GIV, infected and received H. hemistemon extract (100 mg/kg); and GV, infected and received H. hemistemon extract (200 mg/kg). When GIII, GIV, and GV were compared to GII, parasitological analyses displayed highly significant differences in the mean numbers of Cryptosporidium parvum oocysts in the stool between the different groups. GV demonstrated the highest efficacy of 79%. Histopathological analyses displayed improvement in the small intestine and liver pathology in the treated groups (GIII, IV, and V) related to the model (GII), with GV showing the highest efficacy. Moreover, the docking-based study tentatively highlighted the potential of benzoic acid derivatives as lactate dehydrogenase inhibitors. The docked compounds showed the same binding interactions as oxamic acid, where they established H-bond interactions with ARG-109, ASN-140, ASP-168, ARG-171, and HIS-195. To sum up, H. hemistemon is a promising natural therapeutic agent for cryptosporidiosis.

Cryptosporidiosis outbreaks — United States, 2009–2017

Cryptosporidium is a parasite that causes cryptosporidiosis, a profuse, watery diarrhea that can last up to 3 weeks in immunocompetent patients and can lead to life-threatening malnutrition and wasting in immunocompromised patients.* Fecal-oral transmission can occur by ingestion of contaminated recreational water, drinking water, or food, or through contact with infected persons or animals. For the period 2009-2017, public health officials from 40 states and Puerto Rico voluntarily reported 444 cryptosporidiosis outbreaks resulting in 7,465 cases. Exposure to treated recreational water (e.g., in pools and water playgrounds) was associated with 156 (35.1%) outbreaks resulting in 4,232 (56.7%) cases. Other predominant outbreak exposures included contact with cattle (65 outbreaks; 14.6%) and contact with infected persons in child care settings (57; 12.8%). The annual number of reported cryptosporidiosis outbreaks overall increased an average of approximately 13% per year over time. Reversing this trend will require dissemination of prevention messages to discourage swimming or attending child care while ill with diarrhea and encourage hand washing after contact with animals. Prevention and control measures can be optimized by improving understanding of Cryptosporidium transmission through regular analysis of systematically collected epidemiologic and molecular characterization data.

Development of a combined in vitro cell culture – Quantitative PCR assay for evaluating the disinfection performance of pulsed light for treating the waterborne enteroparasite Giardia lamblia

Giardia lamblia is a flagellated protozoan parasite that is recognised as a frequent cause of water-borne disease in humans and animals. We report for the first time on the use of a combined in vitro HCT-8 cell culture-quantitative PCR assay for evaluating the efficacy of using pulsed UV light for treating G. lamblia parasites. Findings showed that current methods that are limited to using vital stains before and after cyst excystation are not appropriate for monitoring or evaluating cyst destruction post PUV-treatments. Use of the human ileocecal HCT-8 cell line was superior to that of the human colon Caco-2 cell line for in vitro culture and determining PUV sensitivity of treated cysts. G. lamblia cysts were also shown to be more resistant to PUV irradiation compared to treating similar numbers of Cryptosporidium parvum oocysts. These observations also show that the use of this HCT-8 cell culture assay may replace use of animal models for determining disinfection performances of PUV for treating both C. parvum and G. lamblia.

A Comparison of Fecal Percent Dry Matter and Number of Cryptosporidium parvum Oocysts Shed to Observational Fecal Consistency Scoring in Dairy Calves

Evaluation of dairy calf feces is often used in research and for clinical decision making to assess severity of diarrhea. However, this has not been validated for agreement between dry matter content and observed fecal consistency. Therefore, a comparison of observed fecal consistency score to fecal percent dry matter and Cryptosporidium parvum oocyst shedding was performed to assess the accuracy of observational scoring as a measure of diarrhea and its association with number of oocysts shed. Fecal samples from 20 dairy calves experimentally infected with C. parvum oocysts were collected daily post-infection and scored on a scale from 1 to 4, with 1 being normal feces to 4 being severe diarrhea. An aliquot of each sample was analyzed for percent dry matter and Cryptosporidium oocyst counts by using immunofluorescent microscopy. Fecal consistency scores of 1, 2, 3, and 4 had median percent dry matter of 20.9, 16.3, 9.6, and 5.8, respectively. Using percent dry matter assessed by fecal consistency scoring were significantly different from each other (P < 0.001). A higher fecal consistency score also was associated with a greater number of Cryptosporidium oocysts shed (P < 0 .0001). Scores of 1, 2, 3, and 4 had median oocyst counts of 0, 0, 1.3 × 10⁶, and 2.8 × 10⁶, respectively. These results suggest that observational scoring is a useful proxy to assess diarrhea in dairy calves.

Report of cryptosporidiosis in gray brocket deer (Mazama gouazoupira) in southern Brazil

The aim of this study was to report the occurrence of cryptosporidiosis in gray brocket deer in Brazil. Fecal samples of a male and a female Mazama gouazoupira kept in captivity in southern Brazil were evaluated. Samples collected at 10-d intervals were analyzed by the centrifugal flotation technique and stained by the modified Ziehl–Neelsen and by the Kinyoun staining methods. The first analysis showed a high number of Cryptosporidium parvum oocysts in the feces, associated with the presence of diarrhea in both animals. However, even without drug therapy, the number of oocysts of the protozoan decreased and the feces returned to normal after 10 days. This is the first report of cryptosporidiosis in M. gouazoupira.

Detection of a Small Number of Cryptosporidium parvum Oocysts by Sugar Floatation and Sugar Centrifugation Methods

Detection rates from the samples including a small number of Cryptosporidium parvum oocysts were compared between the sugar flotation and the sugar centrifugal flotation methods. As the results, the oocysts were detected from 70 and 80 of 100 samples including 6.0x10(2) and 1.0x10(3) oocysts per 1 ml by the flotation method, respectively, whereas from 52 and 53 of the same samples by the centrifugal flotation method. Therefore, it was considered that the flotation method is the most suitable method for the detection from samples including a small number of Cryptosporidium oocysts. It is also suggested that results of the sugar flotation method were reliable for samples including more than 1.0x10(3) oocysts/ml.

Detection of Cryptosporidium parvum in soil extracts.

Epifluorescent microscopy and flow cytometry were used in different combinations with fluorescein isothiocyanate-labeled immunoglobulins M and G3 to estimate the numbers of Cryptosporidium parvum oocysts in soil extracts containing 10 to 10,017 oocysts/ml. No combination had a systematic effect on accuracy or precision. Background debris may have produced overestimates at low oocyst concentrations when flow cytometry was used.

Prophylactic use of decoquinate for infections with Cryptosporidium parvum in experimentally challenged neonatal calves

To evaluate the effect of daily oral administration of decoquinate to neonatal calves experimentally challenged with various numbers of Cryptosporidium parvum oocysts. Clinical trial. 75 calves. Calves were purchased from a commercial dairy during a 5-week period. Calves were housed in individual hutches and fed milk replacer with or without decoquinate (2 mg/kg [0.9 mg/lb per day]). Calves were randomly assigned to treatment and 1 of 5 challenge groups (0, 50, 100, 1000, or 10,000 C. parvum oocysts in 60 mL of saline [0.9% NaCl] solution administered p.o. on the day after arrival). Calves were maintained in the study for as long as 28 days. Calves were clinically assessed for diarrhea and dehydration. Fecal samples were submitted for oocyst enumeration 3 times each week. Treatment did not affect number of days to first watery feces (diarrhea), number of days to first oocyst shedding, or duration of diarrhea or oocyst shedding. Duration of oocyst shedding was significantly associated with challenge dose of oocysts administered to calves and number of days to first oocyst shedding. Duration of diarrhea and number of days to first oocyst shedding were significantly associated with week of arrival and number of days to first watery diarrhea. Daily treatment with decoquinate at the dosage used in this study did not affect oocyst shedding or clinical signs associated with cryptosporidiosis. However, there was an indication that if the number of oocysts calves received could be reduced, then the duration of oocyst shedding and, hence, environmental loading of C. parvum oocysts could be reduced.

Seasonal change in the number of Cryptosporidium parvum oocysts in water samples from the rivers in Hokkaido, Japan, detected by the ferric sulfate flocculation method.

An epidemiological study was carried out in natural water supplies of Hokkaido, one of the largest dairy prefectures in Japan. To investigate the prevalence of Cryptosporidium parvum (C. parvum) oocysts water samples were collected from three rivers in the eastern area of Hokkaido from August 1999 to October 2001, and C. parvum oocysts were collected and purified by the ferric sulfate flocculation method. The oocysts were detected using the immunofluorescent assay test (IFAT) and 4', 6-diamidino-2-phenylindole (DAPI) staining. The seasonal change in the number of oocysts detected was observed. Oocysts increased in numbers from the late summer to the early autumn (from August to November), thereafter, they exhibited a trend to decrease until December, when no oocysts could be detected. The maximum number of oocysts detected in the three rivers was 3.50, 5.00 and 3.33 oocysts/l, respectively. The oocyst density in river water changed in relation to the season in 1999, 2000 and 2001. This report first cleared up the seasonal changes in C. parvum oocysts number in river water.

Number of Cryptosporidium parvum oocysts or Giardia spp cysts shed by dairy calves after natural infection.

To determine the total number of Cryptosporidium parvum oocysts and Giardia spp cysts shed by dairy calves during the period when they are most at risk after natural infection. 478 calves naturally infected with C. parvum and 1,016 calves naturally infected with Giardia spp. Oocysts or cysts were enumerated from fecal specimens. Distribution of number of oocysts or cysts versus age was used to determine the best fitting mathematic function. Number of oocysts or cysts per gram of feces for a given duration of shedding was computed by determining the area under the curve. Total number of oocysts or cysts was calculated by taking the product of the resultant and the expected mass of feces. Intensity of Cparvum oocyst shedding was best described by a second-order polynomial function. Shedding increased from 4 days of age, peaked at day 12, and then decreased. An infected 6-day-old calf would produce 3.89 x 10(10) oocysts until 12 days old. Pattern of shedding of Giardia spp cysts was best described by exponential functions. Intensity of shedding increased from 4 days of age, peaked at day 14, and then decreased. An infected calf would produce 3.8 x 10(7) cysts from day 50 until day 56. The large number of oocysts and cysts shed indicates that shedding by dairy cattle poses a risk for susceptible calves and people. Estimates reported here may be useful to aid in designing cost-effective strategies to manage this risk.

Quantitative polymerase chain (QPCR) reaction using the MIMIC approach to estimate Cryptosporidium parvum oocysts, an intestinal pathogen, in municipal water treatment sludge samples

An accurate estimation of the number of Cryptosporidium parvum oocysts in water treatment plant sludge was determined using the Quantitative Polymerase Chain Reaction (QPCR) method. Approximately 8×106purified viable oocysts were spiked into raw water and treated by conventional water treatment methods. The settled sludge was collected and the DNA extracted. The QPCR Mimic produced two competing products that were 300 and 435 base pairs in size. The log ratio of the products were used in the standard curve to determine a final estimation of oocysts in the sludge sample. The final number of oocysts in the sludge sample was estimated at 258 oocyst per two litres of treated water. This is the first time sludge from a water treatment process has been tested for presence of C. parvum oocysts, which is a known contaminant of drinking water. The QPCR method can be used to test other sludge samples and help estimate the sanitary risks associated with using sludge to fertilize agricultural lands.

Susceptibility and serologic response of healthy adults to reinfection with Cryptosporidium parvum.

Healthy adults are susceptible to infection with small numbers of Cryptosporidium parvum oocysts, resulting in self-limited infection. We investigated if infection of humans with C. parvum is protective 1 year after primary exposure. At 1 year after a primary challenge with 30 to 10(6) oocysts, 19 healthy immunocompetent adults were rechallenged with 500 oocysts and monitored for the development of infection and/or illness. Oocyst excretion was quantitated by direct immunofluorescence with a C. parvum-specific monoclonal antibody, and anti-C. parvum antibodies in serum were detected by an enzyme-linked immunosorbent assay. Fewer subjects shed oocysts after the second exposure (3 of 19; 16%) than after the first exposure (12 of 19; 63%) (P < 0.005). Although the rates of diarrhea were comparable after each of the two exposures, the clinical severity as determined by the mean number of unformed stools passed was lower after reexposure (11.25 versus 8.62; P < 0.05). The number of anti-Cryptosporidium immunoglobulin G and A seroconversions increased after secondary exposure. However, the C. parvum serum antibody response did not correlate with the presence or absence of infection.

The infectivity of Cryptosporidium parvum in healthy volunteers.

Small numbers of Cryptosporidium parvum oocysts can contaminate even treated drinking water, and ingestion of oocysts can cause diarrheal disease in normal as well as immunocompromised hosts. Since the number of organisms necessary to cause infection in humans is unknown, we performed a study to determine the infective dose of the parasite in healthy adults. After providing informed consent, 29 healthy volunteers without evidence of previous C. parvum infection, as determined by the absence of anti-cryptosporidium-specific antibodies, were given a single dose of 30 to 1 million C. parvum oocysts obtained from a calf. They were then monitored for oocyst excretion and clinical illness for eight weeks. Household contacts were monitored for secondary spread. Of the 16 subjects who received an intended dose of 300 or more oocysts, 14 (88 percent) became infected. After a dose of 30 oocysts, one of five subjects (20 percent) became infected, whereas at a dose of 1000 or more oocysts, seven of seven became infected. The median infective dose, calculated by linear regression, was 132 oocysts. Of the 18 subjects who excreted oocysts after the challenge dose, 11 had enteric symptoms and 7 (39 percent) had clinical cryptosporidiosis, consisting of diarrhea plus at least one other enteric symptom. All recovered, and there were no secondary cases of diarrhea among household contacts. In healthy adults with no serologic evidence of past infection with C. parvum, a low dose of C. parvum oocysts is sufficient to cause infection.

Effect of Ultraviolet Disinfection of Drinking Water on the Viability of Cryptosporidium parvum Oocysts

Demineralized water was enriched with a known number of Cryptosporidium parvum oocysts obtained from fresh calf feces, which were purified and exposed to ultraviolet (UV) light (15,000 mW/sec) for different lengths of time. Coccidium-free litters of CD-1 neonatal mice then were inoculated intragastrically with the treated water. Seven days postinoculation mice were killed and C. parvum infection prevalence and intensity determined. In mice inoculated with C. parvum-enriched water that had been exposed to UV light for at least 150 min, no infection occurred.

Threshold of detection of Cryptosporidium oocysts in human stool specimens: evidence for low sensitivity of current diagnostic methods

To determine the minimum number of Cryptosporidium oocysts that can be detected in stool specimens by diagnostic procedures, stool samples seeded with known numbers of Cryptosporidium parvum oocysts were processed by the modified Formalin-ethyl acetate (FEA) stool concentration method. FEA concentrates were subsequently examined by both the modified cold Kinyoun acid-fast (AF) staining and fluorescein-tagged monoclonal antibody (immunofluorescence [IF]) techniques. Oocysts were more easily detected in watery diarrheal stool specimens than they were in formed stool specimens. For watery stool specimens, a 100% detection rate was accomplished at a concentration of 10,000 oocysts per g of stool by both the AF staining and IF techniques. In formed stool specimens, 100% of specimens seeded with 50,000 oocysts per gram of stool were detected by the IF technique, whereas 500,000 oocysts per g of stool were needed for a 100% detection rate by AF staining. Counting of all oocysts on IF slides indicated a mean oocyst loss ranging from 51.2 to 99.6%, depending on the stool consistency as determined by the FEA concentration procedure. Our findings suggest that the most commonly used coprodiagnostic techniques may fail to detect cryptosporidiosis in many immunocompromised and immunocompetent individuals.