3.1 Isolation and characterisation of animal samples
3.1.3 Seasonality, age and spatial distribution of cats shedding T. gondii
Statistical analysis of the age of oocyst-shedding cats and the seasonality of oocyst-shedding was only carried out for the period between 2007 and 2008 and for feline faecal samples submitted by VetMed Labor GmbH since for that period and from this particular laboratory all data sets were obtained for complete analy-sis. For each month, the total number of feline faecal samples received by VetMed Labor GmbH was inquired and correlated to the number of T. gondii positive fae-cal samples as analysed by PCR. The numbers of T. gondii-positive and -negative faecal samples were analysed by the Fisher’s Exact Test for each month and summarised for each quarter (QT).
A total of 18,259 feline faecal samples were received for diagnosis at Vet-Med Labor GmbH. During the whole period, 45/18,259 (0.25%) cats shed T.
gondii oocysts, whereas 33/18,259 (0.18%) cats shed H. hammondi oocysts.
Among all the samples received at VetMed Labor GmbH between 2007 and 2008, the proportion of T. gondii-positive samples (n = 45; 0.25%) differed depending on the submission month and the quarter of the year (Table 14 and Table 15). In the first (January–March), second (April–June), third (July–September) and fourth quarter (October–December), 0.16% (4/2,549), 0.03% (1/2,953), 0.37%
(21/5,734) or 0.27% (19/7,023) of the samples tested positive for T. gondii (Table 15). To compare T. gondii-positive and -negative feline faecal samples quarter wise, Fisher’s Exact Test was applied. The differences were only statistically sig-nificant (Fisher’s Exact Test; P > 0.05) when the 2nd and the 3rd quarter (P = 0.002) and the 2nd and 4th quarter (P = 0.01) were compared (Table 16). When combining several months, statistically significant differences were observed be-tween the 2nd quarter and quarters 1+3+4 as well as between the 3rd quarter and quarters 1+2+4. Following this analysis, the year was divided into two halves ranging from January-June and from July-December. T. gondii positive and nega-tive faecal sample were again analysed for any statistically significant differences.
When the 1st half of the year (January–June) was compared to the 2nd half (July–
December), a significantly higher proportion of samples tested positive for T.
gondii oocysts in the 2nd half of the year (P = 0.005).
Table 14: Total number of samples analysed by VetMed Labor GmbH and number of samples analysed positive for T. gondii or H. hammondi per month.
Total number of samples and number of samples tested.
Month Samples T. gondii positive
(%) H. hammondi positive (%)
January 1,007 1 (0.10) 4 (0.40)
February 829 1 (0.12) 0 (0.00)
March 713 2 (0.28) 1 (0.14)
April 794 0 (0.00) 0 (0.00)
May 688 0 (0.00) 0 (0.00)
June 1,471 1 (0.07) 1 (0.07)
July 1,791 8 (0.45) 1 (0.06)
August 1,841 10 (0.54) 3 (0.16)
September 2,102 3 (0.14) 1 (0.05)
October 2,541 7 (0.28) 8 (0.31)
November 2,488 8 (0.32) 9 (0.36)
December 1,994 4 (0.20) 5 (0.25)
Totals 18,259 45 (0.25) 33 (0.18)
Table 15: Total number of samples analysed by VetMed Labor GmbH and number of samples tested positive for T. gondii or H. hammondi per quarter of the year.
Total number of samples and number of samples tested.
Quarter (QT),
(period) Samples T. gondii
positive (%) H. hammondi positive (%) QT1 (January–March) 2,549 4 (0.16) 5 (0.20) QT2 (April–June) 2,953 1 (0.03) 1 (0.03) QT3 (July–September) 5,734 21 (0.37) 5 (0.09) QT4 (October–December) 7,023 19 (0.27) 22 (0.31)
Totals 18,259 45 (0.25) 33 (0.18)
Table 16: Statistical analysis (Fisher’s Exact Test) of T. gondii oocyst shedding for different quar-ters of the year.
QT=Quarter, Statistically significant differences (P < 0.05, Fisher’s Exact test) are indicated by bold letters.
Comparison T. gondii positive;
T. gondii negative [n] T. gondii positive;
T. gondii negative [n] Significance [P =]
QT1 vs QT2 4; 2,545 1; 2,952 0.1893
QT1 vs QT3 4; 2,545 21; 5,713 0.1304
QT1 vs QT4 4; 2,545 19; 7,004 0.4780
QT2 vs QT3 1; 2,952 21; 5,713 0.0024
QT2 vs QT4 1; 2,952 19; 7,004 0.0128
QT3 vs QT4 21; 5,713 19; 7,004 0.3443
QT1 vs QT2+3+4 4; 2,545 41; 15,669 0.3959
QT2 vs QT1+3+4 1; 2,952 44; 15,262 0.0070
QT3 vs QT1+2+4 21; 5,713 24; 12,501 0.0355
QT4 vs QT1+2+3 19; 7,004 26; 11,210 0.6464
QT1+2 vs QT3+4 5; 5,497 40; 12,717 0.0050
Another aim of the study was to determine if T. gondii oocyst shedding could be correlated to a certain age or age group. The age ranges of the cats from which T. gondii-positive samples were obtained were therefore compared accord-ingly. Three categories of age ranges were formed: group 1) one year and younger, group 2) 1–7 years and group 3) older than seven years. Complete data sets were available for 34 cats. Among these animals, 18/34 (52.94%) positive cats were up to one year old. Sixteen animals (16/34; 47.06%) were older than one year of age.
6/34 (17.65%) of T. gondii-positive cats were between one and seven years old.
Only five T. gondii-positive cats (5/34; 14.71%) were below the age of six months. Ten (10/34; 29.41%) cats were older than seven years and the oldest cat shown to shed T. gondii oocysts was 18 years old. The results are summarised in Figure 7. Samples from ten T. gondii-negative cats were randomly selected per month (i.e. a total number of 190 cats) and included into the statistical analysis.
These samples allowed comparison of T. positive cats with T. gondii-negative cats with regards to (i) the age when they shed oocysts and (ii) the geo-graphical locations where the cat samples had been submitted from. The age dis-tribution among the T. positive cats was similar to that of the T. gondii-negative cats. In the gondii-negative sample 81/190 (42.63%) of the cats were up to one year old, 47/190 (24.74%) cats were 1–7 years old and 62/190 (32.63%) cats were older than seven years old. No statistically significant difference regarding the proportion of cats between T. gondii-positive and T. gondii-negative animals was observed (Fisher’s Exact test; P > 0.05).
It was assumed that cats kept in rural areas were more susceptible to infec-tion with T. gondii and thus more likely to shed oocysts. Cats in those areas were also more likely to roam freely and had thus easier access to infected intermediate host species of T. gondii, such as birds and rodents. It was also more likely that cats in such areas were fed less processed and tinned cat food thus increasing the likelihood of getting infected and to shed oocysts. It was thus hypothesised that cats originating from areas with a high population density were less likely to shed
oocysts than cats from rural areas with a lower population density. To test this hypothesis, the possible influence of human population density on the shedding of T. gondii oocysts of cats was also analysed. If the postal code of the place where the cat sample had been submitted from was available, the respective data set was analysed by comparing the geographical origin of the T. gondii-negative samples with that of the T. gondii-positive samples with regard to human population densi-ty at the district level. T. gondii-negative samples came from districts with a mean human population density of 710.50 (CI: 643.10–777.90) persons per square kil-ometre (km2), whereas T. gondii-positive samples came from districts with a mean population density of 815.80 (CI: 666.20–965.20) persons per km2. The differ-ences in the population density were not statistically significant between the dis-tricts where negative and positive samples had been submitted from (P > 0.05;
Mann-Whitney U Test). Only three out of 44 (6.82%) T. gondii-positive samples came from districts with a population density of less than 100 persons per km2. Twelve positive samples (27.27%) originated from districts with a population den-sity of 100–500 persons per km2, whereas 29 positive samples (65.91%) came from districts with a population density of more than 500 persons per km2 (Figure 8).
Figure 7: Age distribution of cats which shed T. gondii oocysts between June 2007 and December 2008. Distribution is shown as the proportion relative to the total number (n = 34) of positive cats which shed T. gondii in the respective period.
Figure 8: Map of Germany showing where T. gondii-positive and -negative samples (June 2007-December 2008) originated from.
Red dots indicate T. gondii-negative sample; green dots represent T. gondii-positive sample. The size of the dots corresponds to different sample size. Population density (inhabitants/km2) is shown in purple shading.